MXPA00009442A

MXPA00009442A - Serine protease variants having amino acid deletions and substitutions

Info

Publication number: MXPA00009442A
Application number: MXPA/A/2000/009442A
Authority: MX
Inventors: Nelton Rubingh Donn; Elizabeth Ellen Sikorski; Paul Elliott Correa
Original assignee: The Procter&Ampgamble Company
Priority date: 1998-03-26
Filing date: 2000-09-26
Publication date: 2001-07-09

Abstract

The present invention relates to variants of serine proteases having decreased immunogenicity relative to their corresponding wild-type proteases. More particularly, the present invention relates to variants having a modified amino acid sequence of a wild-type amino acid sequence, wherein the modified amino acid sequence comprises a deletion and, optionally, a substitution of one or more specifically identified positions corresponding to subtilisin BPN'. The invention further relates to mutant genes encoding such variants and cleaning and personal care compositions comprising such variants.

Description

SERINE PROTEASE VARIANTS THAT HAVE DELETIONS AND SUBSTITUTIONS OF AMINO ACIDS FIELD OF THE INVENTION The present invention relates to serine protease variants and compositions comprising the variants and having decreased immunogenicity in relation to their corresponding wild-type serine proteases.

BACKGROUND OF THE INVENTION Enzymes are the largest class of proteins that occur naturally. One class of enzyme includes proteases that catalyze the hydrolysis of other proteins. This ability to hydrolyze proteins has been exploited by incorporating proteases that occur naturally and proteases manipulated by genetic engineering in cleaning compositions, particularly those pertinent to laundry applications. In cleaning techniques, the most widely used of these proteases are serine proteases. Most of these serine proteases are produced by bacterial organisms while some are produced by other organisms, such as fungi. See Siezen, Roland J. et al., "Homology Modeling and Protein Engineering Strategy of Subtilases, the Family of Subtilisin-Like Serine Proteases", Protein Engineering, Vol. 4, No. J, pp. 719-737 (1991). Unfortunately, the effectiveness of wild-type proteases in their natural environment often does not translate to the environment of the unnatural cleaning composition. Specifically, the characteristics of the protease such as, for example, thermal stability, pH stability, oxidative stability and substrate specificity are not necessarily optimized by utilizing the enzyme outside the natural environment. Various methods have been employed to alter the wild-type amino acid sequence of serine proteases with the goal of increasing the efficacy of the protease in the wash environment that is not natural. These approaches include the genetic redesign of proteases to increase thermal stability and to improve stability to oxidation under very diverse conditions. However, because such genetically engineered proteases are foreign to mammals, they are potential antigens. As antigens, these proteases produce immunological and allergic responses (collectively described here as immunological responses) in mammals. In fact, sensitization to serine proteases has been observed in environments where humans are regularly exposed to proteases. Such environments include manufacturing facilities, where employees are exposed to proteases through vehicles such as uncontrolled dust or aerosolization. Aerosolization can occur by introducing the protease into the lung, which is the route of exposure of the protease that causes the most dangerous response. Protease sensitization can also occur in the market, where the repeated use of products containing proteases by consumers can cause an allergic reaction. In addition, although genetic engineering has been prominent in the continuing search for more highly effective proteases to be used in laundry applications, genetically engineered proteases have been used to a minimum in personal care compositions and light duty detergents. A major reason for the absence of genetically engineered proteases in products such as, for example, soaps, gels, body washes and shampoos is due to the aforementioned problem of human sensitization leading to undesirable immune responses. Therefore, it would be highly advantageous to provide a personal care composition that would provide the cleaning properties of the proteases manipulated by genetic engineering with minimal provocation of immune responses. One method that leads to the alleviation of the immunological activity of a protease is through the redesign of one or more epitopes of the protease. Epitopes are those regions of amino acids in an antigen that induce an immune response through the binding of antibodies or the presentation of processed antigens to T cells through a major histocompatibility complex protein (MHC). Changes in epitopes can affect its efficiency as an antigen. See Walsh, B. J. and M.E.H. Howden, "A Method for the Detection of IgE Binding Sequences of Allergens Based on a Modification of Epitope Mapping", Journal of Immunological Methods, Vol. 121, pp. 275-280 (1989). The inventors of the present invention have discovered that those serine proteases commonly known as subtilisins, including subtilisin BPN ', have a prominent epitope region at positions 70-84 of amino acids corresponding to BPN'. The inventors of the present invention have genetically redesigned here such subtilisins to remedy the immunogenic properties attributed to this epitope region. In doing so, the inventors of the present invention have discovered subtilisins that induce a diminished immune response but maintain their activity as an efficient cleaning protease. Accordingly, the proteases of the present invention are suitable for use in various types of compositions including, but not limited to laundry, dishwashing, hard surfaces, skin care, hair care, beauty care, oral compositions and compositions for contact lenses.

BRIEF DESCRIPTION OF THE INVENTION The present invention relates to serine protease variants having decreased immunogenicity in relation to their corresponding wild-type proteases. Very particularly, the present invention relates to variants having a modified amino acid sequence of a wild-type amino acid sequence, wherein the modified amino acid sequence comprises a deletion (deletion) and optionally a substitution of one or more positions 70- 84 corresponding to subtilisin BPN '. The invention also relates to mutant genes encoding the variants and cleansing and personal care compositions containing said variants.

DETAILED DESCRIPTION OF THE INVENTION The essential components of the present invention are described below. Also included are non-limiting descriptions of various optional and preferred components useful in the embodiments of the present invention. The present invention may comprise, consist of or consist essentially of any of the required or optional components and / or limitations described herein. All percentages and ratios are calculated by weight unless otherwise indicated. All percentages are calculated based on the total composition unless otherwise indicated. All levels of component or composition are in reference to the active level of that component or composition, and are exclusive of impurity, for example, residual solvents or by-products, which may be present in commercially available sources. All documents referred to herein, including all patents, patent applications and printed publications, are hereby incorporated by reference in their entirety. As used herein, abbreviations will be used to describe amino acids. Table 1 provides a list of abbreviations used in the present invention.

TABLE 1 Definitions As used herein, the term "mutation" refers to alterations in gene sequences and amino acid sequences produced by those gene sequences. The mutations can be deletions, substitutions or additions of amino acid residues to the sequence of wild-type proteins. As used herein, the term "wild type" refers to a protein, specifically a protease, produced by organisms that have not undergone mutation. As used herein, the term "variant" means a protein, specifically a protease, having an amino acid sequence that differs from that of the wild-type protease. As referred to herein, although the variants of the present invention are not limited to those of the subtilisins BPN ', the numbering of all the amino acids is with reference to the amino acid sequence for the subtilisin BPN' which is represented by SEQ ID NO. : 1. The amino acid sequence for subtilisin BPN 'is further described in Wells, JA, E. Ferrari, DJ Henner, D.A. Estell, and E.Y. Chen, Nucleic Acids Research, Vol. II, 7911-7925 (1983), incorporated herein by reference.

Variants of the present invention The inventors of the present invention have discovered an epitope region in serine protease corresponding to positions 70-84 of subtilisin BPN '. The inventors of the present invention have also discovered that one or more deletions and / or amino acid substitutions in the amino acid sequence of a wild-type serine protease provides variants that induce a decreased allergenic response and / or decreased immune response relative to the corresponding wild-type protease resin. As used herein, a variant can be designated by referring to the truncated amino acid positions characterizing the variant, for example, a variant of a serine protease having deletions at positions 70, 75, 76, 77, 78 , 79, 80, 81 and 82 can be designated as 70, 75-82. Similarly, substitutions can be indicated by providing the wild type amino acid residue, followed by the position number, and followed by the substituted amino acid residue to be substituted. Where the substituted amino acid residue can be any natural amino acid allowed in that particular position, the symbol "*" is given. Multiple substitutions that comprise a variant are separated by the "+" symbol. To illustrate this, a substitution of glycine for valine at position 70 is designated GlyJOVal or G70V. A variant comprising both deletions and substitutions is designated by combining the aforementioned designations. For example, an example of a variant having a substitution in both positions 70 and 72, as well as deletions in positions 75-82, is designated as 75-82, G70V + V72A.

The variants of the present invention are variants of serine protease. As used herein, the term "serine protease" means a protease having at least 50%, and preferably 80%, of amino acid sequence identity with the sequence for one or more of a serine protease similar to subtilisin. A discussion related to serine proteases similar to subtilisin and its homologies can be found in Siezen et al., "Homology Modeling and Protein Engineering Strategy of Subtilisases, the Family of Subtilisin-Like Serine Proteases", Protein Engineering, Vol. 4, No. 7, pp. 719-737 (1991). Preferred serine proteases for mutation include subtilisin BPN ', subtilisin Carisberg, subtilisin DY, subtilisin 309, proteinase K and termitase. The serine protease that is most preferred for mutation is subtilisin BPN '. The variants of the present invention are variants of serine proteases having a modified amino acid sequence of a wild-type amino acid sequence, wherein the modified amino acid sequence comprises a deletion of one or more of positions 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83 and 84 (70-84) corresponding to BPN '. Unless you understand a composition for personal care, the variants here are not? J5 - 83 or? J1. Most preferably, the variants of the present invention comprise a deletion of one or more of the positions of 73, 74, 75, 76, 77, 78, 79, 80, 81, 82 and 83 (73-83), most preferably a one or more than 75, 76, 77, 78, 79, 80, 81 and 82 (75-82).

Preferred variants of the present invention include? 70, 75-82; ? 75-82; ? 70, 78, 79; ? 70; ? 75; ? 76; ? 78; ? 79; ? 81 and? 82. The most preferred variants include? 70, 75 -82; ? 75-82; ? 70, 78, 79; ? 70; ? 78; Y ? 79. The variants of the present invention, in addition to one or more deletions, may optionally comprise a substitution of one or more of positions 70-84 corresponding to subtilisin BPN '. Of course, if a deletion has been made in any given position, a substitution can not be made in that position. Substitutions in the epitope region are made by replacing the wild-type amino acid residue with another natural amino acid residue such as that given in Table 1. Tables 2-10 below illustrate non-limiting variants of the present invention. With respect to these tables, when describing the specific mutations the wild type amino acid residue is given first and the corresponding position number is given in the second term. Table 2 outlines preferred individual choice variants. Tables 3-10 delineate preferred variants having one or more deletions and optionally one or more substitutions. To illustrate this, the example in Table 3 reads "LEU 82 GLY 83". This example is intended to illustrate three different variants: one that has deletions at positions 82 and 83 (? 82, 83), one that has a deletion at position 82 and a substitution at 83 (? 82, G83 *), and a that has a substitution in position 82 and a deletion in position 83 (? 83, L82 *).

TABLE 2 Variants of a single deletion VAL S GLY S3 LEU 82 VAL SI GLY te ILE 79 SER 71 A 7? ASN 76 LEU 73 WING 74 10 WING 73 VAL 72 GLY 70 TABLE 3 Double mutation variants LEU 12 GLY 13 15 VAL SI GLY 13 VAL tl LEU 12 GLY 10 GLY 13 GLY 10LEU 12 GLY 10 VAL 11 1LE 7 GLY 13 ULE 79 LEU 12 ILE n VAL 11 20 ILE 79GLY 10 SE * 71 GLY 13 SER 71 LEU 12 SER 71 VAL 1 »lEfc 7S GLY 10 SER 71 BLE 9 ASN 77 GLY 83 ASN 77 LEU 82 ASN 77 VAL 81 ASN 77 GLY 80 ASN 77 ILE 79 ASN 77 SER 78 ASN 76 GLY 83 ASN 76 LEU 82 ASN 76 VAL 81 ASN 76 GLY 80 ASN 76 ÍLE 79 ASN 76 SER 78 ASN 76 ASN 77 LEU 75 GLY 83 LEU 75 LEU 82 LEU 75 VAL SI LEU 75 GLY 80 LEU 75 LE 79 LEU 75 SER 78 LEU 75 ASN 77 LEU 75 ASN 76 WING 74 GLY 83 WING 74 WAY 82 WING 74 WAY WING 74 WAY 80 WING 74 ILE 79 WING 74 WAY 78 WING 74 WED 77 WING 74 WED 76 WED 74 WED 75 WING 73 GLY 83 WING 73 WED 82 WING 73 Wing 81 WING 73 Wing 80 Wing 73 Wing 79 Wing 73 Wing 78 Wing 73 Wing 73 WING 73 Wing 73 WING 73 Wing 73 Wing 73 Wing 73 Wing 74 TABLE 4 Triple mutation variants VAL 81 LEU 82 GLY 83 GLY 80 LEU 82 GLY 83 GLY 80 VAL 81 GLY 83 GLY 80 VAL 81 LEU 82 ILE 79 LEU 82 GLY 83 1LE 79 VAL 81 GLY 83 ILE 79 VAL 1 LEU 82 ILE 79 GLY 80 GLY 83 ILE 79 GLY 80 LEU 82 ILE 79 GLY 80 VAL 81 SER 78 LEU 82 GLY 83 SER 78 VAL 81 GLY 83 SER 78 VAL 81 LEU 82 SER 78 GLY 80 GLY 83 SER 78 GLY 80 LEU 82 SER 78 GLY 80 VAL 81 SER 78 ILE 79 GLY 83 SE 78 ILE 79 LEU 82 SER 78 ILE 79 VAL 81 SER 78 ILE 79 GLY 80 ASN 77 LEU 82 GLY 83 ASN 77 VAL 81 GLY 83 ASN 77 VAL 81 LEU 82 ASN 77 GLY 80 GLY 83 ASN 77 GLY 80 LEU 82 ASN 77 GLY 80 VAL 81 ASN 77 ILE 79 GLY 83 ASN 77 ELE 79 LEU 82 ASN 77 ELE 79 VAL 81 ASN 77 ILE 79 GLY 80 ASN 77 SER 78 GLY 83 ASN 77 SER 78 LEU 82 ASN 7? SER 78 VAL 81 ASN 77 SER 78 GLY 80 ASN 77 SER 78 ILE 79 ASN 76 LEU 82 GLY 83 ASN 76 VAL 81 GLY 83 ASN 76 VAL 1 LEU 82 ASN 76 GLY 80 GLY 83 ASN 76 GLY 80 LEU 82 ASN 76 GLY 80 VAL 81 ASN 76 ULE 79 GLY 83 ASN 76 ELE 79 LEU 82 ASN 76 ÍLE 79 VAL 81 ASN 76 ILE 79 GLY 80 ASN 76 BE 78 GLY 83 ASN 76 BE 78 LEU 82 ASN 76 BE 78 VAL 81 ASN 76 BE 78 GLY 80 ASN 76 BE 78 ILE 79 ASN 76 ASN 77 GLY 83 ASN 76 ASN 77 LEU 82 ASN 76 ASN 77 VAL 81 ASN 76 ASN 77 GLY 80 ASN 76 ASN 7 ILE 79 ASN 76 ASN 77 BE 78 LEU 75 LEU 82 GLY 83 LEU 75 VAL 81 GLY 83 LEU 75 VAL 81 LEU 82 LEU 75 GLY 80 GLY 83 LEU 75 GLY 80 LEU 82 LEU 75 GLY 80 VAL 81 LEU 75 ELE 79 GLY 83 LEU 75 ILE 79 LEU 82 LEU 75 ILE 79 VAL 81 LEU 75 1LE 79 GLY 80 LEU 75 SER 78 GLY 83 LEU 75 SER 78 LEU 82 LEU 75 SER 78 VAL 81 LEU 75 SER 78 GLY 80 LEU 75 SER 78 ÍLE 79 LEU 75 ASN 77 GLY 83 LEU 75 ASN 77 LEU 82 LEU 75 ASN 77 VAL 81 LEU 75 ASN 77 GLY 80 LEU 75 ASN 77 ILE 79 LEU 75 ASN 77 SER 78 LEU 75 ASN 76 GLY 83 LEU 75 ASN 76 LEU 82 LEU 75 ASN 76 VAL 81 LEU 75 ASN 76 GLY 80 LEU 75 ASN 7 ILE 79 LEU 75 ASN 76 SER 78 LEU 75 ASN 76 ASN 77 WING 74 LEU 82 GLY 83 WING 74 VAL 81 GLY 83 WING 74 VAL 81 LEU 82 WING 74 GLY 80 GLY 83 WING 74 GLY 80 LEU 82 WING 74 WED 80 WED 81 WED 74 WED 79 WED 83 WING 74 WED 79 WEST 82 WING 74 WEST 79 WEST 81 WING 74 WEST 79 WEST 80 WING 74 WIT 78 GLY 83 WING 74 SER 78 LEU 82 WING 74 SER 78 VAL 81 WING 74 SER 78 GLY 80 WING 74 SER 7 ILE 79 WING 74 ASN 77 GLY 83 WING 74 ASN 77 LEU 82 WING 74 ASN 77 WAL 81 WING 74 WET 77 WET 80 WING 74 ASN 77 ILE 79 ALA 74 ASN 77 BE 78 WING 74 ASN 76 GLY 83 WING 74 ASN 76 LEU 82 WING 74 ASN 76 VAL 81 WING 74 ASN 76 GLY 80 WING 74 ASN 76 ILE 79 WING 74 ASN 76 WING 78 WING 74 ASN 76 ASN 77 WING 74 LEU 75 GLY 83 WING 74 LEU 75 LEU 82 WING 74 LEU 75 VAL 81 ALA 74 LEU 75 GLY 80 ALA 74 LEU 75? LE 79 ALA 74 LEU 75 SER 78 WING 74 LEU 75 ASN 77 WING 74 LEU 75 ASN 76 WING 73 LEU 82 GLY 83 WING 73 VAL 81 GLY 83 WING 73 VAL 81 LEU 82 ALA 73 GLY 80 GLY 83 ALA 73 GLY 80 LEU 82 ALA 73 GLY 80 VAL 81 ALA 73 ILE 79 GLY 83 WING 73 ILE 79 LEU 82 WING 73 ILE 79 VAL 81 WING 73 ILE 79 GLY 80 WING 73 SER 78 GLY 83 ALA 73 SER 78 LEU 82 ALA 73 SER 78 VAL 81 WING 73 SER 78 GLY 80 WING 73 SER 78 ILE 79 WING 73 ASN 77 GLY 83 WING 73 ASN 77 LEU 82 WING 73 ASN 77 W 81 W 73 W 77 W 80 W 73 ASN 77 ILE 79 ALA 73 ASN 77 SER 78 ALA 73 ASN 76 GLY 83 ALA 73 ASN 76 LEU 82 ALA 73 ASN 76 VAL 81 ALA 73 ASN 76 GLY 80 ALA 73 ASN 76 ILE 79 ALA 73 ASN 76 SER 78 ALA 73 ASN 76 ASN 77 ALA 73 LEU 75 GLY 83 ALA 73 LEU 75 LEU 82 ALA 73 LEU 75 VAL 81 ALA 73 LEU 75 GLY 80 10 ALA 73 LEU 75 ILE 79 ALA 73 LEU 75 SER 78 ALA 73 LEU 75 ASN 77 ALA 73 LEU 75 ASN 76 ALA 73 ALA 74 GLY 83 ALA 73 ALA 74 LEU 82 ALA 73 ALA 74 VAL 81 ALA 73 ALA 74 GLY 80 ALA 73 ALA 74 ILE 79 ALA 73 ALA 74 SER 78 ALA 73 ALA 74 ASN 77 ALA 73 ALA 74 ASN 76 ALA 73 ALA 74 LEU 75 TABLE 5 20 Quadruple mutation variants GLY 80 VAL 81 LEU 82 GLY 83 ILE 79 VAL 81 LEU 82 GLY 83 ILE 79 GLY 80 LEU 82 GLY 83 ILE 79 GLY 80 I VAL 81 GLY 83 ILE 79 GLY 80 VAL 81 LEU 82 SER 78 VAL 81 LEU 82 GLY 83 SER 78 GLY 80 LEU 82 GLY 83 BE 78 GLY 80 VAL 81 GLY 83 BE 78 GLY 80 VAL 81 LEU 82 BE 78 ILE 79 LEU 82 GLY 83 BE 78 ILE 79 I VAL 81 GLY 83 BE 78 ILE 79 I VAL 81 LEU 82 IRE 78 ILE 79 GLY 80 GLY 83 BE 78 1LE 79 GLY 80 LEU 82 SER 7 ILE 79 GLY 80 VAL 81 ASN 77 VAL 81 LEU 82 GLY 83 ASN 77 GLY 80 LEU 82 GLY 83 ASN 77 GLY 80 VAL 81 GLY 83 ASN 77 GLY 80 VAL 81 LEU 82 ASN 77 ILE 79 LEU 82 GLY 83 ASN 77 ILE 79 VAL 81 GLY 83 ASN 77 ILE 79 VAL 81 LEU 82 ASN 77? LE 79 GLY 80 GLY 83 ASN 77 ILE 79 GLY 80 LEU 82 ASN 77 ILE 79 GLY 80 VAL 81 ASN 77 SER 78 LEU 82 GLY 83 ASN 77 SER 78 VAL 81 GLY 83 ASN 77 SER 78 VAL 81 LEU 82 ASN 77 SER 78 GLY 80 GLY 83 ASN 77 SER 78 GLY 80 LEU 82 ASN 77 SER 78 GLY 80 VAL 81 ASN 77 SER 78 ILE 79 GLY 83 ASN 77 SER 78 ILE 79 LEU 82 ASN 77 SER 78 ILE 79 VAL 81 ASN 77 SER 78 ILE 79 GLY 80 ASN 76 VAL 1 LEU 82 GLY 83 ASN 76 GLY 80 LEU 82 GLY 83 ASN 76 GLY 80 VAL 81 GLY 83 ASN 76 GLY 80 VAL 81 LEU 82 ASN 76 ILE 79 LEU 82 GLY 83 ASN 76 ILE 79 VAL 81 GLY 83 ASN 76 1LE 79 VAL 81 LEU 82 ASN 76 ILE 79 GLY 80 GLY 83 ASN 76 ILE 79 GLY 80 LEU 82 ASN 76 ILE 79 GLY 80 VAL 81 ASN 76 SER 78 LEU 82 GLY 83 ASN 76 SER 78 VAL 81 GLY 83 ASN 76 SER 78 VAL 1 LEU 82 ASN 76 SER 78 GLY 80 GLY 83 ASN 76 SER 78 GLY 80 LEU 82 ASN 76 SER 78 GLY 80 VAL 81 ASN 76 SER 78 ILE 79 GLY 83 ASN 76 SER 78 ILE 79 LEU 82 ASN 76 SER 78 ILE 79 VAL 81 ASN 76 SER 78 ILE 79 GLY 80 ASN 76 ASN 77 LEU 82 GLY 83 ASN 76 ASN 77 VAL 81 GLY 83 ASN 76 ASN 77 VAL 81 LEU 82 ASN 76 ASN 77 GLY 80 GLY 83 ASN 76 ASN 77 GLY 80 LEU 82 ASN 76 ASN 77 GLY 80 VAL 81 ASN 76 ASN 77 ÍLE 79 GLY 83 ASN 76 ASN 771LE 79 LEU 82 ASN 76 ASN 77 ILE 79 VAL 81 ASN 76 ASN 77 ILE 79 GLY 80 ASN 76 ASN 77 SER 78 GLY 83 ASN 76 ASN 77 SER 78 LEU 82 ASN 76 ASN 77 SER 78 VAL 81 ASN 76 ASN 77 SER 78 GLY 80 ASN 76 ASN 77 SER 78 ÍLE 79 LEU 75 VAL 81 LEU 82 GLY 83 LEU 75 GLY 80 LEU 82 GLY 83 LEU 75 GLY 80 VAL 81 GLY 83 LEU 75 GLY 80 VAL 81 LEU 82 LEU 75 ILE 79 LEU 82 GLY 83 LEU 75 ILE 79 VAL 81 GLY 83 LEU 75 ILE 79 VAL 81 LEU 82 LEU 75 ILE 79 GLY'80 GLY 83 LEU 75 ILE 79 GLY 80 LEU 82 LEU 75 ILE 79 GLY 80 VAL 85 LEU 75 SER 78 LEU 82 GLY 83 LEU 75 SER 78 VAL 81 GLY 83 LEU 75 BE 78 VAL 81 LEU 82 LEU 75 BE 78 GLY 80 GLY 83 LEU 75 BE 78 GLY 80 LEU 82 LEU 75 BE 78 GLY 80 VAL 81 LEU 75 BE 78 ÍLE 79 GLY 83 LEU 75 SER 78 ILE 79 LEU 82 LEU 75 SER 7 ILE 79 VAL 81 LEU 75 SER 78 ILE 79 GLY 80 LEU 75 ASN 77 LEU 82 GLY 83 LEU 75 ASN 77 VAL 81 GLY 83 LEU 75 ASN 77 VAL 81 LEU 82 LEU 75 ASN 77 GLY 80 GLY 83 LEU 75 ASN 77 GLY 80 LEU 82 LEU 75 ASN 77 GLY 80 VAL 81 LEU 75 ASN 77 ILE 79 GLY 83 LEU 75 ASN 77 ILE 79 LEU 82 LEU 75 ASN 7 ILE 79 VAL 81 LEU 75 ASN 77 ILE 79 GLY 80 LEU 75 ASN 77 SER 78 GLY 83 LEU 75 ASN 77 SER 78 LEU 82 LEU 75 ASN 77 SER 78 VAL 81 LEU 75 ASN 77 SER 78 GLY 80 LEU 75 ASN 77 SER 78 ILE 79 LEU 75 ASN 76 LEU 82 GLY 83 LEU 75 ASN 76 VAL 81 GLY 83 LEU 75 ASN 76 VAL 81 LEU 82 LEU 75 ASN 76 GLY 80 GLY 83 LEU 75 ASN 76 GLY 80 LEU 82 LEU 75 ASN 76 GLY 80 VAL 81 LEU 75 ASN 76 ILE 79 GLY 83 LEU 75 ASN 76 1LE 79 LEU 82 LEU 75 ASN 76 ÍLE 79 VAL 81 LEU 75 ASN 76 ILE 79 GLY 80 LEU 75 ASN 76 SER 78 GLY 83 LEU 75 ASN 76 SER 78 LEU 82 LEU 75 ASN 76 SER 78 VAL 8! LEU 75 ASN 76 SER 78 GLY 80 LEU 75 ASN 76 SER 78 ILE 79 LEU 75 ASN 76 ASN 77 GLY 83 LEU 75 ASN 76 ASN 77 LEU 82 LEU 75 ASN 76 ASN 77 VAL 81 LEU 75 ASN 76 ASN 77 GLY 80 LEU 75 ASN 76 ASN 77 ÍLE 79 LEU 75 ASN 76 ASN 77 SER 78 ALA 74 VAL 81 LEU 82 GLY 83 WING 74 GLY 80 LEU 82 GLY 83 WING 74 GLY 80 VAL 81 GLY 83 WING 74 GLY 80 VAL 81 LEU 82 WING 74 1LE 79 LEU 82 GLY 83 WING 74 ILE 79 VAL 1 GLY 83 WING 74 JLE 79 VAL 81 LEU 82 WING 74 ELE 79 GLY 80 GLY 83 WING 74 ILE 79 GLY 80 LEU 82 WING 74 1LE 79 GLY 80 VAL 81 WING 74 SER 78 EU 82 GLY 83 WING 74 SER 78 VAL 81 GLY 83 WING 74 SER 78 VAL 81 LEU 82 WING 74 SER 78 GLY 80 GLY 83 WING 74 SER 78 GLY 80 LEU 82 WING 74 SER 78 GLY 80 VAL 81 WING 74 SER 78? LE 79 GLY 83 WING 74 SER 78 1LE 79 LEU 82 WING 74 SER 78 ÍLE 79 VAL 81 WING 74 SER 78 ÍLE 79 GLY 80 WING 74 ASN 77 LEU 82 GLY 83 WING 74 ASN 77 VAL 81 GLY 83 WING 74 ASN 77 VAL 81 LEU 82 WING 74 ASN 77 GLY 80 GLY 83 WING 74 ASN 77 GLY 80 LEU 82 WING 74 ASN 77 GLY 80 VAL 81 WING 74 ASN 77 ILE 79 GLY 83 WING 74 ASN 7 ILE 79 LEU 82 WING 74 ASN 77? LE 79 VAL 81 WING 74 ASN 77 ILE 79 GLY 80 WING 74 ASN 77 SER 78 GLY 83 WING 74 ASN 77 SER 78 LEU 82 WING 74 ASN 77 SER 78 VAL 81 WING 74 ASN 77 SER 78 GLY 80 WING 74 ASN 77 SER 78 ILE 79 WING 74 ASN 76 LEU 82 GLY 83 WING 74 ASN 76 WEAVE 81 GLY 83 WING 74 ASN 76 VAL 81 LEU 82 WING 74 ASN 76 GLY 80 GLY 83 WING 74 ASN 76 GLY 80 LEU 82 WING 74 ASN 76 GLY 80 VAL 81 WING 74 ASN 7 ILE 79 GLY 83 WING 74 ASN 76 ILE 79 LEU 82 WING 74 ASN 76 1LE 79 VAL 81 ALA 74 ASN 76 ILE 79 GLY 80 WING 74 ASN 76 SER 78 GLY 83 WING 74 ASN 76 SER 78 LEU 82 WING 74 ASN 76 SER 78 VAL 81 WING 74 ASN 76 SER 78 GLY 80 WING 74 ASN 76 SER 78 ILE 79 WING 74 ASN 76 ASN 77 GLY 83 WING 74 ASN 76 ASN 77 WED 82 WED 74 WED 76 ASN 76 WED 77 WED 74 WED 74 WED 76 WED 76 WED 80 WED 74 WED 76 WED 76 WED 76 WED 74 WED 75 WED 83 WED 74 LEU 75 VAL 81 GLY 83 WING 74 LEU 75 VAL 81 LEU 82 WING 74 LEU 75 GLY 80 GLY 83 WING 74 LEU 75 GLY 80 LEU 82 WING 74 LEU 75 GLY 80 VAL 81 Wing 74 LEU 75 ILE 79 GLY 83 WING 74 LEU 75? LE 79 LEU 82 WING 74 LEU 75 ILE 79 VAL 81 WING 74 LEU 75 ILE 79 GLY 80 WING 74 LEU 75 SER 78 GLY 83 WING 74 LEU 75 SER 78 LEU 82 WING 74 LEU 75 SER 78 VAL 81 WATER 74 LEU 75 SER 78 GLY 80 WING 74 LEU 75 SER 78 ILE 79 WING 74 LEU 75 ASN 77 OLY 83 WING 74 LEU 75 ASN 77 LEU 82 WING 74 LEU 75 ASN 77 W 81 W 74 W 75 W 75 W 74 W 74 W 75 W 75 W 75 W 77 W 75 W 75 W 75 W 75 W 75 W 75 W 76 W 83 W 74 W 75 W 76 W 82 ALA 74 LEU 75 ASN 76 VAL 81 ALA 74 LEU 75 ASN 76 GLY 80 WING 74 LEU 75 ASN 76 WEA 79 WING 74 LEU 75 ASN 76 Wing 78 Wing 74 LEU 75 ASN 76 Wing 77 WING 73 Wing 81 LEE 82 GLY 83 WING 73 GLY 80 LEU 82 GLY 83 WING 73 GLY 80 WET 81 GLY 83 WING 73 GLY 80 VAL 81 LEU 82 WING 73 BLE 79 LEU 82 GLY 83 ALA 73 BLE 79 VAL 81 GLY 83 WING 73 ILE 79 VAL 81 LEU 82 WING 73 ILE 79 GLY 80 GLY 83 WING 73 ILE 79 GLY 80 LEU 82 WING 73 ILE 79 GLY 80 VAL 83 Wing 73 Wing 78 LEU 82 GLY 83 WING 73 SER 78 VAL 81 GLY 83 WING 73 SER 78 VAL 81 LEU 82 WING 73 SER 78 GLY 80 GLY 83 WAND 73 SER 78 GLY 80 LEU 82 WAND 73 SER 78 GLY 80 VAL 81 WAND 73 SER 78 ILE 79 GLY 83 WING 73 SER 78 ILE 79 LEU 82 ALA 73 SER 781LE 79 VAL 81 ALA 73 SER 78 ILE 79 GLY 80 WING 73 ASN 77 LEU 82 GLY 83 WING 73 ASN 77 VAL 81 GLY 83 WING 73 ASN 77 VAL 81 LEU 82 WING 73 ASN 77 GLY 80 GLY 83 ALA 73 ASN 77 GLY 80 LEU 82 ALA 73 ASN 77 GLY 80 VAL 81 ALA 73 ASN 77 ILE 79 GLY 83 ALA 73 ASN 77 ILE 79 LEU 82 ALA 73 ASN 77.ILE 79 VAL 81 ALA 73 ASN 77 ILE 79 GLY 80 ALA 73 ASN 77 SER 78 GLY 83 ALA 73 ASN 77 SER 78 LEU 82 ALA 73 ASN 77 SER 78 VAL 81 ALA 73 ASN 77 SER 78 GLY 80 ALA 73 ASN 77 SER 78 ILE 79 ALA 73 ASN 76 LEU 82 GLY 83 WING 73 ASN 76 VAL 81 GLY 83 WING 73 ASN 76 VAL 81 LEU 82 WING 73 ASN 76 GLY 80 GLY 83 WING 73 ASN 76 GLY 80 LEU 82 ALA 73 ASN 76 GLY 80 VAL 81 ALA 73 ASN 76 ILE 79 GLY 83 ALA 73 ASN 76 ILE 79 LEU 82 ALA 73 ASN 7 ILE 79 VAL 1 ALA 73 ASN 76 ILE 79 GLY 80 ALA 73 ASN 76 SER 78 GLY 83 ALA 73 ASN 76 SER 78 LEU 82 ALA 73 ASN 76 SER 78 VAL 81 WING 73 ASN 76 SER 78 GLY 80 WING 73 ASN 76 SER 78 ILE 79 WING 73 ASN 76 ASN 77 GLY 83 WING 73 ASN 76 ASN 77 LEU 82 WING 73 ASN 76 ASN 77 W 81 W 73 W 76 ASN 77 GLY 80 ALA 73 ASN 76 ASN 77 ILE 79 ALA 73 ASN 76 ASN 77 SER 78 ALA 73 LEU 75 LEU 82 GLY 83 ALA 73 LEU 75 VAL 81 GLY 83 ALA 73 LEU 75 VAL 81 LEU 82 ALA 73 LEU 75 GLY 80 GLY 83 ALA 73 LEU 75 GLY 80 LEU 82 ALA 73 LEU 75 GLY 80 VAL 81 ALA 73 LEU 75 ILE 79 GLY 83 ALA 73 LEU 75 ILE 79 LEU 82 ALA 73 LEU 75 ILE 79 VAL 81 ALA 73 LEU 75 ILE 79 GLY 80 ALA 73 LEU 75 SER 78 GLY 83 ALA 73 LEU 75 SER 78 LEU 82 ALA 73 LEU 75 SER 78 VAL 81 ALA 73 LEU 75 SER 78 GLY 80 ALA 73 LEU 75 SER 78 ILE 79 WING 73 LEU 75 ASN 77 GLY 83 WING 73 LEU 75 ASN 77 LEU 82 WING 73 LEU 75 ASN 77 VAL 8! ALA 73 LEU 75 ASN 77 GLY 80 ALA 73 LEU 75 ASN 77 ILE 79 ALA 73 LEU 75 ASN 77 SER 78 ALA 73 LEU 75 ASN 76 GLY 83 ALA 73 LEU 75 ASN 76 LEU 82 ALA 73 LEU 75 ASN 76 VAL 81 ALA 73 LEU 75 ASN 76 GLY 80 ALA 73 LEU 75 ASN 76 BLE 79 ALA 73 LEU 75 ASN 76 BE 78 3 i ALA 73 LEU 75 ASN 76 ASN 77 ALA 73 ALA 74 LEU 82 GLY 83 ALA 73 ALA 74 VAL 81 GLY 83 ALA 73 ALA 74 VAL 81 LEU 82 ALA 73 ALA 74 GLY 80 GLY 83 ALA 73 WING 74 GLY 80 LEU 82 WING 73 WING 74 Wing 80 Wing 81 Wing 73 Wing 74 Wing 74 Wing 79 Wing 83 Wing 73 Wing 74 Wing 79 Wing 82 Wing 73 Wing 74 Wing 79 Wing 81 Wing 73 Wing 74 Wing 79 Wing 80 Wing 73 Wing 74 SER 78 GLY 83 WING 73 WING 74 SER 78 LEU 82 WING 73 WING 74 SER 78 VAL 81 WING 73 WING 74 Wing 78 WED 80 WING 73 WING 74 WEST 78 WED 79 WED 73 WING 74 WED 74 WED 83 WED 73 WING 73 WED 74 WED 77 WED 75 WED 73 WED 74 WED 74 WED 77 WED 73 WED 73 WED 74 WED 77 WED 80 WING 73 ALA 74 ASN 77 ILE 79 ALA 73 ALA 74 ASN 77 SER 78 WING 73 WING 74 ASN 76 WED 83 WING 73 WING 74 WED 76 WED 82 WED 73 WING 73 WED 74 WED 76 WAND 76 WING 73 WING 74 WED 76 WEST 80 WING 73 WING 76 WED 76 73 ALA 74 ASN 76 ASN 77 ALA 73 ALA 74 LEU 75 GLY 83 ALA 73 ALA 74 LEU 75 LEU 82 ALA 73 ALA 74 LEU 75 VAL 81 ALA 73 ALA 74 LEU 75 GLY 80 ALA 73 ALA 74 LEU 75 ILE 79 ALA 73 ALA 74 LEU 75 SER 78 ALA 73 ALA 74 LEU 75 ASN 77 ALA 73 ALA 74 LEU 75 ASN 76 TABLE 6 Five-fold mutation variants 1LE 79 GLY 80 VAL 81 LEU 82 GLY 83 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 BE 78 ILE 79 GLY 80 VAL 81 LEU 82 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 ASN 77 ILE 79 VAL 81 LEU 82 GLY 83 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 ASN 77 BE 78 GLY 80 LEU 82 GLY 83 ASN 77 BE 78 GLY 80 VAL 81 GLY 83 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 ASN 77 SER 78 ILE 79 GLY 80 IOL 81 ASN 76 IOL 80 IOL 81 LEU 82 GLY 83 ASN 76 ILE 79 VAL 81 LEU 82 GLY 83 ASN 76 ILE 79 GLY 80 LEU 82 GLY 83 ASN 76 ILE 79 GLY 80 VAL 81 GLY 83 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 ASN 76 BE 78 VAL 81 LEU 82 GLY 83 ASN 76 BE 78 GLY 80 LEU 82 GLY 83 ASN 76 BE 78 GLY 80 VAL 81 GLY 83 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 ASN 76 SER 78 ILE 79 LEU 82 GLY 83 ASN 76 SER 78 1LE 79 VAL 81 GLY 83 ASN 76 SER 78? LE 79 VAL 81 LEU 82 ASN 76 SER 78 ILE 79 GLY 80 GLY 83 ASN 76 BE 78 ILE 79 GLY SO LEU 82 ASN 76 BE 78 ILE 79 GLY 80 VAL 81 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 ASN 76 ASN 77 ING 79 ING 82 GLY 83 ASN 76 ASN 77 IGB 79 IOL 79 GLY 83 ASN 76 ASN 77 ILE 79 IOL 79 LEU 82 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 ASN 76 ASN 77 BE 78 GLY 80 LEU 82 ASN 76 ASN 77 BE 78 GLY 80 VAL 81 ASN 76 ASN 77 BE 78 ILE 79 GLY 83 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 ASN 76 ASN 77 SER 78 ULE 79 GLY 80 LEU 75 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ILE 79 VAL 81 LEU 82 GLY 83 LEU 75 ILE 79 GLY 80 LEU 82 GLY 83 LEU 75 ILE 79 GLY 80 VAL 81 GLY 83 LEU 75 ILE 79 GLY 80 VAL 81 LEU 82 LEU 75 SER 78 VAL 81 LEU 82 GLY 83 LEU 75 SER 78 GLY 80 LEU 82 GLY 83 LEU 75 SER 78 GLY 80 VAL 81 GLY 83 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 LEU 75 SER 78 ILE 79 LEU 82 GLY 83 LEU 75 SER 78 1LE 79 VAL 81 GLY 83 LEU 75 SER 78 ÍLE 79 VAL 81 LEU 82 LEU 75 SER 78? LE 79 GLY 80 GLY 83 LEU 75 SER 78 ÍLE 79 GLY 80 LEU 82 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 LEU 75 ASN 77 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 GLY 80 LEU 82 GLY 83 LEU 75 ASN 77 GLY 80 VAL 81 GLY 83 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 LEU 75 ASN 77? LE 79 LEU 82 GLY 83 LEU 75 ASN 77 ILE 79 VAL 81 GLY 83 LEU 75 ASN 77? LE 79 VAL 81 LEU 82 LEU 75 ASN 77 ILE 79 GLY 80 GLY 83 LEU 75 ASN 77 ILE 79 GLY 80 LEU 82 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 LEU 75 ASN 77 SER 78 LEU 82 GLY 83 LEU 75 ASN 77 SER 78 VAL 81 GLY 83 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 LEU 75 ASN 77 SER 78 GLY 80 GLY 83 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 75 ASN 77 SER 78 ILE 79 GLY 83 LEU 75 ASN 77 SER 78 ILE 79 LEU 82 LEU 75 ASN 77 SER 78? LE 79 VAL 81 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 75 ASN 76 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 ILE 79 LEU 82 GLY 83 LEU 75 ASN 76 ILE 79 VAL 81 GLY 83 LEU 75 ASN 76 ILE 79 VAL 81 LEU 82 LEU 75 ASN 76 ILE 79 GLY 80 GLY 83 LEU 75 ASN 76 ILE 79 GLY 80 LEU 82 LEU 75 ASN 76 ILE 79 GLY 80 VAL 81 LEU 75 ASN 76 BE 78 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 VAL 81 GLY 83 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 LEU 75 ASN 76 SER 78 GLY 80 GLY 83 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 75 ASN 76 SER 78 ILE 79 GLY 83 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 LEU 75 ASN 76 SER 78 1LE 79 GLY 80 LEU 75 ASN 76 ASN 77 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 GLY 80 GLY 83 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 75 ASN 76 ASN 77 ILE 79 GLY 83 LEU 75 ASN 76 ASN 77 ILE 79 LEU 82 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 75 ASN 76 ASN 77 SER 78 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 75 ASN 76 ASN 77 SER 78? LE 79 WING 74 GLY 80 VAL 81 LEU 82 GLY 83 WING 74? LE 79 VAL 81 LEU 82 GLY 83 WING 74 ELE 79 GLY 80 LEU 82 GLY 83 WING 74 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 ILE 79 GLY 80 VAL 81 LEU 82 WING 74 SER 78 VAL 81 LEU 82 GLY 83 WING 74 SER 78 GLY 80 LEU 82 GLY 83 WING 74 SER 78 GLY 80 VAL 81 GLY 83 WING 74 SER 78 GLY 80 VAL 81 LEU 82 WING 74 SER 78 ILE 79 LEU 82 GLY 83 WING 74 SER 78 ILE 79 VAL 81 GLY 83 WING 74 SER 78 ILE 79 VAL 81 LEU 82 WING 74 SER 78 1LE 79 GLY 80 GLY 83 WING 74 SER 78 ILE 79 GLY 80 LEU 82 WING 74 SER 78 ILE 79 GLY 80 VAL 81 WING 74 ASN 77 VAL 81 LEU 82 GLY 83 WING 74 ASN 77 GLY 80 LEU 82 GLY 83 WING 74 ASN 77 GLY 80 VAL 81 GLY 83 WING 74 ASN 77 GLY 80 VAL 81 LEU 82 WING 74 ASN 77 ILE 79 LEU 82 GLY 83 WING 74 ASN 77 WEA 79 W 81 WED 83 WED 74 WED 77 ILE 79 W 81 WED 82 WED 74 WED 77 ILE 79 WED 80 GLY 83 ALA 74 ASN 77 ILE 79 GLY 80 LEU 82 WING 74 ASN 77 ILE 79 GLY 80 VAL 81 WING 74 ASN 77 SER 78 LEU 82 GLY 83 WING 74 ASN 77 SER 78 VAL 81 GLY 83 WING 74 ASN 77 SER 78 VAL 81 LEU 82 WING 74 ASN 77 BE 78 GLY 80 GLY 83 WING 74 ASN 77 BE 78 GLY 80 LEU 82 WING 74 ASN 77 BE 78 GLY 80 WAL 81 WING 74 ASN 77 SER 78 ILE 79 GLY 83 WING 74 ASN 77 SER 78 ILE 79 LEU 82 ALA 74 ASN 77 SER 78 ILE 79 VAL 81 ALA 74 ASN 77 SER 78 ILE 79 GLY 80 WING 74 ASN 76 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 ILE 79 LEU 82 GLY 83 ALA 74 ASN 76 ILE 79 VAL 81 GLY 83 WING 74 ASN 76? LE 79 VAL 81 LEU 82 WING 74 ASN 76 ILE 79 GLY 80 GLY 83 WING 74 ASN 76 ILE 79 GLY 80 LEU 82 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 WING 74 ASN 76 SER 78 LEU 82 GLY 83 WING 74 ASN 76 SER 78 VAL 81 GLY 83 WING 74 ASN 76 SER 78 VAL 81 LEU 82 WING 74 ASN 76 BE 78 GLY 80 GLY 83 WING 74 ASN 76 BE 78 GLY 80 LEU 82 WING 74 ASN 76 BE 78 GLY 80 WAL 81 WING 74 ASN 76 BE 78 ILE 79 GLY 83 WING 74 ASN 76 BE 78 ILE 79 LEU 82 ALA 74 ASN 76 SER 78 ILE 79 VAL 81 WING 74 ASN 76 SER 78 ILE 79 GLY 80 WING 74 ASN 76 ASN 77 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 VAL 81 GLY 83 WING 74 ASN 76 ASN 77 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 GLY 80 GLY 83 WING 74 ASN 76 ASN 77 GLY 80 LEU 82 WING 74 ASN 76 ASN 77 GLY 80 VAL 81 WING 74 ASN 76 ASN 77 ELE 79 GLY 83 WING 74 ASN 76 ASN 77 ILE 79 LEU 82 ALA 74 ASN 76 ASN 77 ILE 79 VAL 81 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 WING 74 ASN 76 ASN 77 SER 78 GLY 83 WING 74 ASN 76 ASN 77 SER 78 LEU 82 WING 74 ASN 76 ASN 77 SER 78 W 81 W 74 W 76 76 77 80 WING 74 ASN 76 ASN 77 SER 78 ILE 79 WING 74 LEU 75 VAL 81 LEU 82 GLY 83 WING 74 LEE 75 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ELE 79 LEU 82 GLY 83 WING 74 LEU 75 ILE 79 VAL 81 GLY 83 ALA 74 LEU 75 ELE 79 VAL 81 LEU 82 WING 74 LEU 75 ILE 79 GLY 80 GLY 83 WING 74 LEU 75 LE 79 GLY 80 LEU 82 WING 74 LEU 75 ILE 79 GLY 80 VAL 81 WING 74 LEU 75 SER 78 LEU 82 GLY 83 ALA 74 LEU 75 SER 78 VAL 81 GLY 83 WING 74 LEU 75 SER 78 VAL 81 LEU 82 WING 74 LEU 75 SER. 78 GLY 80 GLY 83 WING 74 LEU 75 SER 78 GLY 80 LEU 82 WING 74 LEU 75 SER 78 GLY 80 VAL 81 WING 74 LEU 75 SER 78 ILE 79 GLY 83 WING 74 LEU 75 SER 78 1LE 79 LEU 82 WING 74 LEU 75 SER 78 ILE 79 VAL 81 WING 74 LEU 75 SER 78? LE 79 GLY 80 WING 74 LEU 75 ASN 77 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 VAL 81 LEU 82 WING 74 LEU 75 ASN 77 GLY 80 GLY 83 WING 74 LEU 75 ASN 77 GLY 80 LEU 82 WING 74 LEU 75 ASN 77 GLY 80 VAL 81 Wing 74 LEU 75 ASN 77 ILE 79 GLY 83 WING 74 LEU 75 ASN 77 ILE 79 LEU 82 WING 74 LEU 75 ASN 77 ÍLE 79 VAL 81 ALA 74 LEU 75 ASN 77 ILE 79 GLY 80 ALA 74 LEU 75 ASN 77 SER 78 GLY 83 WING 74 LEU 75 ASN 77 SER 78 LEU 82 WING 74 LEU 75 ASN 77 SER 78 VAL 81 WING 74 LEU 75 ASN 77 SER 78 GLY 80 WING 74 LEU 75 ASN 77 SER 78 ÍLE 79 WING 74 LEU 75 ASN 76 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 VAL 1 GLY 83 WING 74 LEU 75 ASN 76 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 GLY 80 LEU 82 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 Wing 74 LEU 75 ASN 7 ILE 79 GLY 83 WING 74 LEU 75 ASN 76 ELE 79 LEU 82 ALA 74 LEU 75 ASN 7 ILE 79 VAL 81 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 WING 74 LEU 75 ASN 76 SER 78 GLY 83 ALA 74 LEU 75 ASN 76 SER 78 LEU 82 WING 74 LEU 75 ASN 76 SER 78 VAL 81 Wing 74 LEU 75 ASN 76 SER 78 GLY 80 WING 74 LEU 75 ASN 76 SER 78 1LE 79 WING 74 LEU 75 ASN 76 ASN 77 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 VAL 81 ALA 74 LEU 75 ASN 76 ASN 77 GLY 80 ALA 74 LEU 75 ASN 76 ASN 77 ILE 79 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ALA 73 GLY 80 VAL 81 LEU 82 GLY 83 WING 73? LE 79 VAL 81 LEU 82 GLY 83 WING 73 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 SER 78 VAL 81 LEU 82 GLY 83 WING 73 SER 78 GLY 80 LEU 82 GLY 83 WAND 73 SER 78 GLY 80 VAL 81 GLY 83 WAND 73 SER 78 GLY 80 VAL 81 LEU 82 ALA 73 SER 78 ILE 79 LEU 82 GLY 83 WING 73 SER 78 ILE 79 VAL 81 GLY 83 WING 73 SER 78 ILE 79 VAL 81 LEU 82 WING 73 SER 78 LE 79 GLY 80 GLY 83 WING 73 SER 78 ELE 79 GLY 80 LEU 82 ALA 73 SER 78 ILE 79 GLY 80 VAL 81 ALA 73 ASN 77 VAL 81 LEU 82 GLY 83 WING 73 ASN 77 GLY 80 LEU 82 GLY 83 ALA 73 ASN 77 GLY 80 VAL 81 GLY 83 ALA 73 ASN 77 GLY 80 VAL 81 LEU 82 ALA 73 ASN 77 ILE 79 LEU 82 GLY 83 ALA 73 ASN 77 ILE 79 VAL 81 GLY 83 ALA 73 ASN 77 ILE 79 VAL 81 LEU 82 ALA 73 ASN 77 ILE 79 GLY 80 GLY 83 WING 73 ASN 77? LE 79 GLY 80 LEU 82 WING 73 ASN 77 ELE 79 GLY 80 VAL 81 ALA 73 ASN 77 SER 78 LEU 82 GLY 83 ALA 73 ASN 77 SER 78 VAL 81 GLY 83 WING 73 ASN 77 SER 78 VAL 81 LEU 82 WING 73 ASN 77 SER 78 GLY 80 GLY 83 WING 73 ASN 77 SER 78 GLY 80 LEU 82 ALA 73 ASN 77 SER 78 GLY 80 VAL 81 ALA 73 ASN 77 SER 78 ILE 79 GLY 83 ALA 73 ASN 77 SER 78 ILE 79 LEU 82 ALA 73 ASN 77 SER 78 1LE 79 VAL 81 ALA 73 ASN 77 SER 78 ILE 79 GLY 80 ALA 73 ASN 76 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 GLY 80 LEU 82 GLY 83 WING 73 ASN 76 GLY 80 VAL 81 GLY 83 WING 73 ASN 76 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 ÍLE 79 LEU 82 GLY 83 ALA 73 ASN 76 ILE 79 VAL 81 GLY 83 ALA 73 ASN 76 ILE 79 VAL 81 LEU 82 ALA 73 ASN 76 ILE 79 GLY 80 GLY 83 ALA 73 ASN 76 1 LE 79 GLY 80 LEU 82 ALA 73 ASN 76 ILE 79 GLY 80 VAL 81 ALA 73 ASN 76 SER 78 LEU 82 GLY 83 ALA 73 ASN 76 SER 78 VAL 81 GLY 83 WING 73 ASN 76 SER 78 VAL 81 LEU 82 WING 73 ASN 76 SER 78 GLY 80 GLY 83 WING 73 ASN 76 SER 78 GLY 80 LEU 82 WING 73 ASN 76 SER 78 GLY 80 VAL 81 ALA 73 ASN 76 SER 78 ILE 79 GLY 83 ALA 73 ASN 76 SER 78 ILE 79 LEU 82 ALA 73 ASN 76 SER 78 ILE 79 VAL 81 ALA 73 ASN 76 SER 78 1LE 79 GLY 80 WING 73 ASN 76 ASN 77 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 GLY 80 GLY 83 ALA 73 ASN 76 ASN 77 GLY 80 LEU 82 ALA 73 ASN 76 ASN 77 GLY 80 VAL 81 WING 73 ASN 76 ASN 77 ÍLE 79 GLY 83 ALA 73 ASN 76 ASN 77 ILE 79 GLE 80 Wing 73 Wing 73 Wing 76 Wing 76 Wing 77 Wing 79 Wing 73 Wing 73 Wing 73 Wing 76 Wing 76 Wing 79 Wing 73 Wing 73 Wing 73 Wing 73 Wing 76 82 ALA 73 ASN 76 ASN 77 SER 78 VAL 81 ALA 73 ASN 76 ASN 77 SER 78 GLY 80 ALA 73 ASN 76 ASN 77 SER 78? LE 79 ALA 73 LEU 75 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 GLY 80 VAL 81 GLY 83 ALA 73 LEU 75 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 ILE 79 LEU 82 GLY 83 ALA 73 LEU 75 ILE 79 VAL 81 GLY 83 ALA 73 LEU 75? LE 79 VAL 81 LEU 82 ALA 73 LEU 75? LE 79 GLY 80 GLY 83 ALA 73 LEU 75 ILE 79 GLY 80 LEU 82 ALA 73 LEU 75 ILE 79 GLY 80 VAL 81 ALA 73 LEU 75 SER 78 LEU 82 GLY 83 ALA 73 LEU 75 SER 78 VAL 81 GLY 83 ALA 73 LEU 75 SER 78 VAL 81 LEU 82 ALA 73 LEU 75 SER 78 GLY 80 GLY 83 ALA 73 LEU 75 SER 78 GLY 80 LEU 82 ALA 73 LEU 75 SER 78 GLY 80 VAL 81 ALA 73 LEU 75 SER 78 ILE 79 GLY 83 ALA 73 LEU 75 SER 78 ÍLE 79 LEU 82 ALA 73 LEU 75 SER 78 ILE 79 VAL 81 ALA 73 LEU 75 SER 78 ILE 79 GLY 80 ALA 73 LEU 75 ASN 77 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 GLY 80 GLY 83 ALA 73 LEU 75 ASN 77 GLY 80 LEU 82 ALA 73 LEU 75 ASN 77 GLY 80 VAL 81 ALA 73 LEU 75 ASN 77 ILE 79 GLY 83 ALA 73 LEU 75 ASN 77 ILE 79 LEU 82 ALA 73 LEU 75 ASN 7 ILE 79 VAL 81 ALA 73 LEU 75 ASN 77 ILE 79 GLY 80 ALA 73 LEU 75 ASN 77 SER 78 GLY 83 ALA 73 LEU 75 ASN 77 SER 78 LEU 82 ALA 73 LEU 75 ASN 77 SER 78 VAL 81 ALA 73 LEU 75 ASN 77 SER 78 GLY 80 ALA 73 LEU 75 ASN 77 SER 78 ILE 79 ALA 73 LEU 75 ASN 76 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 VAL 81 GLY 83 ALA 73 LEU 75 ASN 76 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 GLY 80 GLY 83 ALA 73 LEU 75 ASN 76 GLY 80 LEU 82 ALA 73 LEU 75 ASN 76 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 ILE 79 GLY 83 ALA 73 LEU 75 ASN 76 ILE 79 LEE 82 LEA 73 LEU 75 ASN 76 ILE 79 VAL 81 ALA 73 LEU 75 ASN 76 ILE 79 GLY 80 ALA 73 LEU 75 ASN 76 SER 78 GLY 83 ALA 73 LEU 75 ASN 76 SER 78 LEU 82 ALA 73 LEU 75 ASN 76 SER 78 VAL 81 ALA 73 LEU 75 ASN 76 SER 78 GLY 80 ALA 73 LEU 75 ASN 76 SER 78 ILE 79 ALA 73 LEU 75 ASN 76 ASN 77 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 GLY 80 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 ALA 73 LEU 75 ASN 76 ASN 77 BE 78 ALA 73 ALA 74 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 ILE 79 LEU 82 GLY 83 ALA 73 ALA 74? LE 79 VAL 81 GLY 83 ALA 73 ALA 74 ILE 79 VAL 81 LEU 82 ALA 73 ALA 74 ILE 79 GLY 80 GLY 80 GLY 83 WING 73 WING 74 WED 79 GLY 80 LEU 82 WING 73 WING 74 ILE 79 GLY 80 WAL 81 WING 73 WING 74 Wing 78 Wing 82 Wing 83 Wing 73 Wing 74 Wing 78 Wing 81 WALL 83 ALA 73 ALA 74 SER 78 VAL 81 LEU 82 ALA 73 ALA 74 BE 78 GLY 80 GLY 83 ALA 73 ALA 74 BE 78 GLY 80 LEU 82 ALA 73 ALA 74 BE 78 GLY 80 VAL 81 ALA 73 ALA 74 BE 78 ILE 79 GLY 83 ALA 73 WING 74 SER 78 ILE 79 LEU 82 WING 73 WING 74 SER 78 1LE 79 VAL 8! WING 73 WING 74 SER 78 1LE 79 GLY 80 WING 73 WING 74 ASN 77 LEU 82 GLY 83 WING 73 WING 74 WET 77 WET 81 WET 83 WING 73 WING 74 WET 77 W 81 W 79 WING 73 WING 74 ASN 77 GLY 80 LEU 82 WING 73 WING 74 ASN 77 GLY 80 VAL 81 WING 73 WING 74 ASN 771LE 79 GLY 83 ALA 73 ALA 74 ASN 77 LE 79 LEU 82 ALA 73 ALA 74 ASN 77 ILE 79 VAL 81 ALA 73 ALA 74 ASN 77 ILE 79 GLY 80 ALA 73 ALA 74 ASN 77 SER 78 GLY 83 ALA 73 ALA 74 ASN 77 SER 78 LEU 82 ALA 73 ALA 74 ASN 77 SER 78 VAL 81 ALA 73 ALA 74 ASN 77 SER 78 GLY 80 ALA 73 ALA 74 ASN 77 SER 78 1LE 79 ALA 73 WING 74 ASN 76 LEU 82 WING 83 WING 73 WING 74 WED 76 WAND 76 WAND 81 WAY 83 WING 73 WING 74 WAND 76 WAND 80 GLY 83 WING 73 WING 73 WING 76 WEST 80 WEST 82 ALA 73 ALA 74 ASN 76 GLY 80 VAL 81 WING 73 WING 74 ASN 76 ILE 79 GLY 83 WING 73 WING 74 ASN 76 1LE 79 LEU 82 WING 73 WING 74 WED 76 WED 79 WED 79 WED 73 WED 74 WED 74 WED 79 WED 80 WED 73 WING 74 WED 76 WAND 76 WAND 83 WING 73 WING 73 WING 74 WAND 76 WAND 78 WAY 82 WING 73 WING 74 WAND 76 WAND 76 VAL 81 WING 73 WING 74 ASN 76 SER 78 GLY 80 WING 73 WING 74 WED 74 ASN 76 SER 78 ILE 79 WING 73 WING 74 WED 76 WAND 76 WAND 83 WING 73 WING 73 WING 74 WWN 76 WED 76 WED 82 WED 73 WED 74 WED 76 WED 76 VAL 81 WING 73 WING 74 ASN 76 ASN 77 WING 80 WING 73 WING 74 WED 74 WED 76 WED 76 WED 79 WED 73 WED 74 WED 74 WED 76 WED 76 WED 73 WED 73 WED 74 WED 75 LEU 75 WED 83 WED 73 WED 73 WED 74 WED 75 WED 81 GLY 83 ALA 73 ALA 74 LEU 75 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 ILE 79 GLY 83 ALA 73 ALA 74 LEU 75 ILE 79 LEU 82 ALA 73 ALA 74 LEU 75 ILE 79 VAL 81 ALA 73 ALA 74 LEU 75 ILE 79 GLY 80 ALA 73 ALA 74 LEU 75 SER 78 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 LEU 82 ALA 73 ALA 74 LEU 75 SER 78 VAL 81 WING 73 WING 74 LEU 75 SER 78 GLY 80 WING 73 WING 74 LEU 75 SER 78 ILE 79 ALA 73 ALA 74 LEU 75 ASN 77 ALM 77 ALM 73 ALA 73 ALA 74 LEU 75 ASN 77 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 VAL 81 ALA 73 ALA 74 LEU 75 ASN 77 GLY 80 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 WING 73 WING 74 LEU 75 ASN 76 GLY 83 WING 73 WING 74 LEU 75 ASN 76 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 VAL 81 ALA 73 ALA 74 LEU 75 ASN 76 GLY 80 ALA 73 ALA 74 LEU 75 ASN 76 ALS 79 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 TABLE 7 Sextuple mutation variants BE 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ASN 77 BE 78 GLY 80 VAL 81 LEU 82 GLY 83 ASN 77 BE 78 ILE 79 VAL 81 LEU 82 GLY 83 ASN 77 BE 78 1LE 79 GLY 80 LEU 82 GLY 83 ASN 77 BE 78 ILE 79 GLY 80 VAL 81 GLY 83 ASN 77 BE 78 ILE 79 GLY 80 I VAL 81 LEU 82 ASN 76. { LE 79 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 ASN 76 SER 78 1LE 79 GLY 80 VAL 81 LEU 82 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 ASN 7 ILE 79 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 ASN 76 ASN 77? LE 79 GLY 80 VAL 81 GLY 83 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 ASN 76 ASN 77 BE 78 GLY 80 VAL 81 GLY 83 ASN 76 ASN 77 BE 78 GLY 80 VAL 81 LEU 82 ASN 76 ASN 77 BE 78 ILE 79 LEU 82 GLY 83 ASN 76 ASN 77 BE 78 ÍLE 79 VAL 81 GLY 83 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 75 ÍLE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 SER 78 1LE 79 VAL 81 LEU 82 GLY 83 LEU 75 SER 78 1LE 79 GLY 80 LEU 82 GLY 83 LEU 75 SER 78 1LE 79 GLY 80 VAL 81 GLY 83 LEU 75 SER 78 1LE 79 GLY 80 VAL 81 LEU 82 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 ILE 79 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 I LE 79 GLY 80 LEU 82 GLY 83 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 BE 78 GLY 80 LEU 82 GLY 83 LEU 75 ASN 77 BE 78 GLY 80 VAL 81 GLY 83 LEU 75 ASN 77 BE 78 GLY 80 VAL 81 LEU 82 LEU 75 ASN 77 BE 78 ILE 79 LEU 82 GLY 83 LEU 75 ASN 77 BE 78 ILE 79 VAL 81 GLY 83 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ELE 79 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ILE 79 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 ILE 79 GLY 80 I VAL 81 GLY 83 LEU 75 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 ÍLE 79 VAL 81 GLY 83 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 GLY 83 LEU 75 ASN 76 BE 78 ELE 79 GLY 80 LEU 82 LEU 75 ASN 76 BE 78 ELE 79 GLY 80 VAL 81 LEU 75 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 ILE 79 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 LEU 75 ASN 76 ASN 77 1LE 79 GLY 80 VAL 81 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 GLY SO LEU 82 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 75 ASN 76 ASN 77 SER 78 1LE 79 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 75 ASN 76 ASN 77 SER 78 1LE 79 GLY 80 WING 74 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 74 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 74 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 77? LE 79 VAL 81 LEU 82 GLY 83 WING 74 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 ALA 74 ASN 77? LE 79 GLY 80 VAL 81 GLY 83 WING 74 ASN 77 ELE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 74 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 ALÁ 74 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 74 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 74 ASN 77 SER 78 - LE 79 LEU 82 GLY 83 WING 74 ASN 77 SER 78 - LE 79 VAL 81 GLY 83 WING 74 ASN 77 SER 78 ELE 79 VAL 81 LEU 82 WING 74 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 74 ASN 77 SER 78 - LE 79 GLY 80 LEU 82 WING 74 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 WING 74 ASN 76 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 JLE 79 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ILE 79 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 SER 78? LE 79 LEU 82 GLY 83 WING 74 ASN 76 SER 78 ILE 79 VAL 81 GLY 83 WING 74 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 WING 74 ASN 76 SER 78 ÍLE 79 GLY 80 GLY 83 WING 74 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 WING 74 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 WING 74 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 76 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 ILE 79 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 ILE 79 VAL 81 GLY 83 WING 74 ASN 76 ASN 77 1LE 79 VAL 83 LEU 82 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 WING 74 ASN 76 ASN 77 1LE 79 GLY 80 LEU 82 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 WING 74 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 WING 74 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 WING 74 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 WING 74 ASN 76 ASN 77 SER 78 ÍLE 79 GLY 83 WING 74 ASN 76 ASN 77 SER 78? LE 79 LEU 82 WING 74 W ASN 76 ASN 77 SER 78 WEL 79 W 81 W 74 W 76 W 76 W 79 W 74 W 74 W 80 W 81 W 81 W 83 ALA 74 LEU 75 ILE 79 ILE 79 LEU 82 GLY 83 ALA 74 LEU 75 ILE 79 GLY 80 LEU 82 GLY 83 ALA 74 LEU 75 ILE 79 GLY 80 I VAL 81 GLY 83 I ALA 74 LEU 75 1LE 79 GLY 80 VAL 81 LEU 82 ALA 74 LEU 75 SER 78 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 SER 78 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 SER 78 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 BE 78 ILE 79 LEU 82 GLY 83 WING 74 LEU 75 SER 78 ILE 79 VAL 81 GLY 83 WING 74 LEU 75 SER 78 ILE 79 VAL 81 LEU 82 WING 74 LEU 75 SER 78 ILE 79 GLY 80 GLY 83 WING 74 LEU 75 SER 78 ILE 79 GLY 80 LEU 82 WING 74 LEU 75 SER 78 1LE 79 GLY 80 VAL 81 WING 74 LEU 75 ASN 77 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 77 ILE 79 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 ILE 79 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 ILE 79 VAL 81 LEU 82 WING 74 LEU 75 ASN 77 ILE 79 GLY 80 GLY 83 WING 74 LEU 75 ASN 77 ILE 79 GLY 80 LEU 82 ALA 74 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 WING 74 LEU 75 ASN 77 SER 78 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 ALA 74 LEU 75 ASN 77 SER 78 GLY 80 GLY 83 WING 74 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 WING 74 LEU 75 ASN 77 SER 78 ÍLE 79 GLY 83 ALA 74 LEU 75 ASN 77 SER 78 ILE 79 LEU 82 WING 74 LEU 75 ASN 77 SER 78 ELE 79 VAL 81 Wing 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 WING 74 LEU 75 ASN 76 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 JLE 79 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ILE 79 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 ILE 79 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 LEU 82 WING 74 LEU 75 ASN 76 ÍLE 79 GLY 80 VAL 81 WING 74 LEU 75 ASN 76 SER 78 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 SER 78 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 WING 74 LEU 75 ASN 76 SER 78 ÍLE 79 GLY 83 WING 74 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 WING 74 LEU 75 ASN 76 SER 78 ÍLE 79 VAL 81 Wing 74 LEU 75 ASN 76 SER 78? LE 79 GLY 80 WING 74 LEU 75 ASN 76 ASN 77 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 VAL 81 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 LEU 82 WING 74 LEU 75 ASN 76 ASN 77? LE 79 VAL 81 ALA 74 LEU 75 ASN 76 ASN 77? LE 79 GLY 80 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 WING 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 WING 74 LEU 75 ASN 76 ASN 77 SER 78 1LE 79 WING 73 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 BE 78 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 73 SER 78? LE 79 GLY 80 LEU 82 GLY 83 WING 73 SER 78 ÍLE 79 GLY 80 VAL 81 GLY 83 WING 73 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 ASN 77? LE 79 VAL 81 LEU 82 GLY 83 WING 73 ASN 77 WEA 79 GLY 80 LEU 82 GLY 83 WING 73 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 73 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 73 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 WING 73 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 WING 73 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 73 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 ALA 73 ASN 77 BE 78 ELE 79 GLY 80 LEU 82 ALA 73 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 WING 73 ASN 76 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 ASN 76? LE 79 GLY 80 VAL 81 GLY 83 WING 73 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 WING 73 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 WING 73 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 SER 78 ILE 79 LEU 82 GLY 83 WING 73 ASN 76 SER 78 ILE 79 VAL 81 GLY 83 WING 73 ASN 76 SER 78 ELE 79 VAL 81 LEU 82 WING 73 ASN 76 SER 78 ILE 79 GLY 80 GLY 83 WING 73 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 ALA 73 ASN 76 BE 78 ELE 79 GLY 80 VAL 81 ALA 73 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 ILE 79 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 ILE 79 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 WING 73 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 WING 73 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 WING 73 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 WING 73 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 WING 73 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 ALA 73 ASN 76 ASN 77 SER 78 ILE 79 GLY 83 WING 73 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 WING 73 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 ALA 73 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 ALA 73 LEU 75 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 1LE 79 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 SER 78 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 SER 78 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 SER 78 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 SER 78 ILE 79 LEU 82 GLY 83 WING 73 LEU 75 SER 78 ILE 79 VAL 81 GLY 83 WING 73 LEU 75 SER 78 ILE 79 VAL 81 LEU 82 ALA 73 LEU 75 SER 78 DLE 79 GLY 80 GLY 83 WING 73 LEU 75 SER 78 ILE 79 GLY 80 LEU 82 WING 73 LEU 75 SER 78 ÍLE 79 GLY 80 VAL 81 ALA 73 LEU 75 ASN 77 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 ILE 79 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 ILE 79 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 ELE 79 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 ILE 79 GLY 80 GLY 83 WING 73 LEU 75 ASN 77 ILE 79 GLY 80 LEU 82 WING 73 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 Wing 73 LEU 75 ASN 77 SER 78 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 SER 78 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 SER 78 GLY 80 GLY 83 WING 73 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 WING 73 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 Wing 73 LEU 75 ASN 77 SER 78 ? LE 79 GLY 83 WING 73 LEU 75 ASN 77 SER 78 ILE 79 LEU 82 WING 73 LEU 75 ASN 77 SER 78? LE 79 VAL 81 Wing 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 ALA 73 LEU 75 ASN 76 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 ILE 79 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ILE 79 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 ELE 79 VAL 81 LEU 82 WING 73 LEU 75 ASN 76? LE 79 GLY 80 GLY 83 ALA 73 LEU 75 ASN 76 ILE 79 GLY 80 LEU 82 ALA 73 LEU 75 ASN 76 ELE 79 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 SER 78 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 SER 78 VAL 81 GLY 83 ALA 73 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 SER 78 GLY 80 GLY 83 WING 73 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 WING 73 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 83 WING 73 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 WING 73 LEU 75 ASN 76 SER 78 ELE 79 VAL 81 Wing 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 WING 73 LEU 75 ASN 76 ASN 77 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 ASN 77 GLY 80 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 WING 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 WING 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 WING 73 WING 74 GLY 80 W 81 W 82 W 83 ALA 73 ALA 74 ILE 79 VAL 81 LEU 82 GLY 83 ALA 73 WING 74 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ALA 74 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 WING 74 SER 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 SER 78 GLY 80 VAL 81 LEU 82 WING 73 WING 74 SER 78 ILE 79 LEU 82 GLY 83 WING 73 WING 74 Wing 78 ILE 79 W 81 WED 83 WING 73 WED 74 WEL 78 WEL 79 W 81 W 82 W 73 W 74 W 78 W 78 W 79 W 73 W 73 W 74 W 78 W 80 W 80 W 82 W 73 WING 74 SER 78 ILE 79 GLY 80 VAL 81 WING 73 WING 74 WAND 77 WAND 81 WAND 83 WING 83 WING 73 WING 74 WEST 77 WEST 80 WEST 83 WED 83 WING 73 WED 74 WEST 77 WEST 80 WEST 81 WALL 83 WING 73 WING 74 WAND 77 WEST 80 W 81 W 82 WED 73 WING 73 WED 74 WED 77 WIL 79 LEU 82 WED 83 WING 73 WED 74 WED 77 WED 79 WED 83 WED 83 WED 73 WED 74 WED 77 WED 79 W 81 WED 82 WED 73 WING 74 ASN 77 WEA 79 W 79 W 79 W 79 W 73 W 74 W 74 W 77 W 77 W 74 W 83 W 73 W 79 W 73 W 73 W 73 W 73 74 ASN 77 SER 78 VAL 81 GLY 83 WING 73 WING 74 ASN 77 SER 78 VAL 81 LEU 82 WING 73 WING 74 WAND 77 WAND 78 GLY 80 WAND 83 WING 73 WING 74 WAND 77 WAND 78 GLY 80 WEST 82 WING 73 WING 74 WWN 77 SER 78 GLY 80 VAL 81 WING 73 WING 74 ASN 77 SER 78 ILE 79 GLY 83 WING 73 WING 74 ASN 77 SER 78 ILE 79 LEU 82 WING 73 WING 74 ASN 77 SER 78 ILE 79 VAL 81 Wing 73 WING 74 ASN 77 SER 78 ILE 79 GLY 80 WING 73 WING 74 ASN 76 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 GLY 80 LEU 82 GLY 83 WING 73 WING 74 WED 76 WED 80 WAT 81 WED 83 WING 73 WING 74 WED 74 WED 76 WED 80 WALL 81 LEU 82 WING 73 WING 74 WED 76 WED 79 LEU 82 GLY 83 WING 73 WED 74 WED 76 WED 79 WAL 81 WED 83 WING 73 ALA 74 ASN 76 ILE 79 VAL 81 LEU 82 ALA 73 WING 74 ASN 76 ILE 79 GLY 80 GLY 83 WING 73 WING 74 ASN 76 LLE 79 GLY 80 LEU 82 WING 73 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 WING 73 WING 74 WAND 76 WAND 78 WAY 82 WAY 83 WING 73 WING 74 WAND 76 WAND 76 WAND 81 WAND 83 WING 73 WING 74 WAND 76 WAND 78 WAND 81 WET 75 WING 73 WING 73 WED 74 WEST 76 73 WING 74 ASN 76 SER 78 GLY 80 LEU 82 WING 73 WING 74 ASN 76 SER 78 GLY 80 W 81 W 73 W W 74 W T 76 76 W 79 W 83 WING 73 WING 74 WED 76 SER 78 ILE 79 LEU 82 WING 73 WING 74 WED 74 WED 76 WAND 78 W 79 W 81 W 73 W 74 W 76 W 76 W 76 W 76 W 76 W 76 73 WING 74 ASN 76 ASN 77 VAL 81 GLY 83 WING 73 ALA 74 ASN 76 ASN 77 VAL 81 LEU 82 ALA 73 ALA 74 ASN 76 ASN 77 GLY 80 GLY 83 ALA 73 ALA 74 ASN 76 ASN 77 GLY 80 LEU 82 ALA 73 ALA 74 ASN 76 ASN 77 GLY 80 VAL 81 ALA 73 ALA 74 ASN 76 ASN 771LE 79 GLY 83 WING 73 WING 74 ASN 76 ASN 77 ILE 79 LEU 82 WING 73 WING 74 ASN 76 ASN 77 ILE 79 Wing 81 WING 73 WING 74 Wing 76 Wing 76 Wing 79 Wing 80 Wing 80 Wing 73 Wing 74 WED 76 ASN 77 SER 78 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 VAL 81 WING 73 WING 74 ASN 76 ASN 77 SER 78 GLY 80 WING 73 WING 74 ASN 76 ASN 77 BE 78 ILE 79 WING 73 WING 74 LEU 75 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 GLY 80 VAL 81 GLY 83 WING 73 WING 74 WED 75 WED 80 WED 81 LEU 82 WING 73 WING 74 WED 75 LEE 75 ILE 79 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ILE 79 W 81 WED 83 WED 73 WED 74 WED 75 W 75 W 81 W 81 W 73 ALA 74 LEU 75 ILE 79 GLY 80 GLY 83 WING 73 WING 74 LEU 75 ILE 79 GLY SO LEU 82. WING 73 WING 74 LEU 75 ILE 79 GLY 80 VAL 81 WING 73 WING 74 LEU 75 WELD 78 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 VAL 81 GLY 83 WING 73 WING 74 LEU 75 SER 78 VAL 81 LEU 82 WING 73 WING 74 LEU 75 SER 78 GLY 80 GLY 83 WING 73 WING 74 LEU 75 SER 78 GLY 80 LEU 82 WING 73 ALA 74 LEU 75 SER 78 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 SER 78 ELE 79 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 LEU 82 ALA 73 ALA 74 LEU 75 SER 78? LE 79 VAL 81 WING 73 WING 74 WATER 75 WEST 78 WEST 79 WEST 80 WING 73 WING 74 WED 74 LEW 75 WEST 75 LEN 82 WEL 83 WING 83 WING 73 WED 74 LEU 75 WAND 77 W 81 WED 83 WED 73 WED 74 LEU 75 WEST 77 W 81 WED 73 ALA 74 LEU 75 ASN 77 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 ASN 77 ELE 79 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 1LE 79 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 VAL 81 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 GLY 80 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 LEU 82 WING 73 WING 74 LEU 75 ASN 77 SER 78 VAL 81 Wing 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 WING 73 WING 74 LEU 75 ASN 77 SER 78 ILE 79 WING 73 WING 74 LEU 75 ASN 76 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 76 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 ASN 76 ELE 79 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ILE 79 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 ELE 79 VAL 81 ALA 73 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 VAL 81 ALA 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 WING 73 WING 74 LEU 75 ASN 76 SER 78 ELE 79 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 83 WING 73 ALA 74 LEU 75 ASN 76 ASN 77 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 VAL 81 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 GLY 80 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 ÍLE 79 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 TABLE 8 Sevenfold mutation variants ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77 SER 78 LLE 79 VAL 81 LEU 82 GLY 83 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 LEU 75 SER 78? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 BE 78 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 LEU 75 ASN 77 SER 78 ÍLE 79 GLY 80 VAL 8 i GLY 83 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 1LE 79 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 ELE 79 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77? LE 79 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 1LE 79 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 BE 78 ELE 79 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 BE 78 ELE 79 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 ELE 79 VAL 81 LEU 82 LEU 75 ASN 76 ASN 77 BE 78 ELE 79 GLY 80 GLY 83 LEU 75 ASN 76 ASN 77 BE 78 ILE 79 GLY 80 LEU 82 LEU 75 ASN 76 ASN 77 BE 78 ELE 79 GLY 80 VAL 81 WING 74 BE 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 77 ILE 79 GLY 80 VAL 8] LEU 82 GLY 83 WING 74 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 77 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 74 ASN 77 SER 78 ÍLE 79 GLY 80 LEU 82 GLY 33 WING 74 ASN 77 SER 78 ILE 79 GLY 80 VAL SI GLY 83 WING 74 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 SER 78 1LE 79 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 SER 78 1LE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77? LE 79 GLY SO LEU 82 GLY 83 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 ASN 77? LE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 SER 78 ÍLE 79 LEU 82 GLY 83 ALA 74 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 ALA 74 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 WING 74 ASN 76 ASN 77 SER 78? LE 79 GLY 80 VAL 81 WING 74 LEU 75 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 SER 78 GLY 80 VAL SI LEU 82 GLY 83 WING 74 LEU 75 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 ALA 74 LEU 75 SER 78 ELE 79 GLY 80 LEU 82 GLY 83 ALA 74 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 SER 78 1LE 79 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 ÍLE 79 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 77? LE 79 GLY 80 VAL 81 LEU 82 ALA 74 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 WING 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ILE 79 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 76? LE 79 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 ALA 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 ALA 74 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 SER 78 ÍLE 79 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 SER 78 ELE 79 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 WING 74 LEU 75 ASN 76 SER 78 ELE 79 GLY 80 VAL 81 WING 74 LEU 75 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 ASN 77? LE 79 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 - LE 79 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 ELE 79 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 WING 74 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 Wing 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 WING 73 SER 78 1LE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 ASN 77 ILE 79 GLY 80 VAL 8! LEU 82 GLY 83 WING 73 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 ASN 77 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 ALA 73 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 ASN 77 SER 78 ELE 79 GLY 80 VAL 81 GLY 83 WING 73 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 ILE 79 GLY 80 VAL SI LEU 82 GLY 83 WING 73 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 SER 78 1LE 79 GLY 80 LEU 82 GLY 83 WING 73 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 ALA 73 ASN 76 SER 78? LE 79 GLY 80 VAL 81 LEU 82 ALA 73 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 ELE 79 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 WING 73 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 WING 73 ASN 76 ASN 77 SER 78 LLE 79 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 73 ASN 76 ASN 77 BE 78 ILE 79 GLY 80 LEU 82 WING 73 ASN 76 ASN 77 BE 78 1LE 79 GLY 80 VAL 81 WING 73 LEU 75 LLE 79 GLY 80 VAL 8! LEU 82 GLY 83 WING 73 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 SER 78 ILE 79 VAL 8! LEU 82 GLY 83 WING 73 LEU 75 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 GLY 83- WING 73 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 I LE 79 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 WING 73 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 ALA 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 1LE 79 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 7 ILE. 79 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 GLY 83 ALA 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 ALA 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 VAL 8! LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 ÍLE 79 VAL 81 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77? LE 79 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ÍLE 79 VAL 81 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 WING 73 WING 74 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 SER 78 BLE 79 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 SER 78 ÍLE 79 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 ASN 77? LE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 ALÁ 73 ALA 74 ASN 77 ELE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 77 SER 78 ÍLE 79 LEU 82 GLY 83 WING 73 WING 74 ASN 77 SER 78 ILE 79 VAL 81 GLY S3 WING 73 WING 74 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 73 WING 74 ASN 77 SER 78 ÍLE 79 GLY 80 GLY 83 WING 73 WING 74 ASN 77 SER 78 LE 79 GLY 80 LEU 82 WING 73 WING 74 WAND 77 WAND 78 WEST 79 WEST 80 W 81 WED 73 WING 74 WAND 76 WEST 80 W 81 W 82 W 83 W 73 W W 74 W 76 76 81 LEU 82 GLY 83 ALA 73 WING 74 ASN 76? LE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ALA 74 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 WING 74 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 76 SER 78? LE 79 LEU 82 GLY 83 WING 73 WING 74 ASN 76 SER 78 ILE 79 VAL 81 GLY 83 WING 73 WING 74 AS N 76 SER 78 ILE 79 VAL 81 LEU 82 WING 73 WING 74 ASN 76 SER 78 ILE 79 GLY 80 GLY 83 WING 73 WING 74 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 WING 73 WING 74 ASN 76 SER 78 1LE 79 GLY 80 VAL 81 WING 73 WING 74 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 76 Wing 77 WING 79 W 82 WED 83 WING 73 WING 74 WED 76 WED 76 WED 79 W 81 WED 83 WED 73 WED 74 WED 76 WED 77 WED 79 W 81 WED 82 WING 73 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 WING 73 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 WING 73 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 WING 73 WING 74 ASN 76 ASN 77 BE 78 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL SI WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 JLE 79 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 ALA 73 ALA 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 WING 73 WING 74 LEU 75 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ILE 79 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75? LE 79 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ILE 79 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 SER 78 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 ÍLE 79 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 SER 78? LE 79 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 SER 78? LE 79 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 ASN 77 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77? LE 79 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ELE 79 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 77 ILE 79 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 77 SER 78 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ÍLE 79 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ILE 79 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ÍLE 79 VAL 81 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ÍLE 79 GLY 80 WING 73 WING 74 LEU 75 ASN 76 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 ÍLE 79 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ILE 79 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 1LE 79 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 ELE 79 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 1LE 79 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 I LE 79 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 76 SER 78 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 1LE 79 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 ÍLE 79 VAL 81 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 1LE 79 GLY 80 WING 73 WING 74 LEU 75 ASN 76 ASN 77 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 VAL 8 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 ÍLE 79 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77? LE 79 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 ÍLE 79 VAL 81 WING 73 WING 74 LEE 75 ASN 76 ASN 77? LE 79 GLY 80 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY SO WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 TABLE 9 Eightfold mutation variants ASN 76 ASN 77 BE 78 ELE 79 GLY 80 VAL 8! LEU 82 GLY 83 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 771LE 79 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 8! LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 LEU 75 ASN 76 ASN 77 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 WING 74 ASN 77 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 1LE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 ÍLE 79 GLY 80 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 SER 78 1LE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 ELE 79 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78? LE 79 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 SER 78 1LE 79 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 77 SER 78 ELE 7 9 GLY 80 VAL 81 LEU 82 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78 1LE 79 GLY 80 VAL SI GLY 83 ALA 74 LEU 75 ASN 76 SER 78 ÍLE 79 GLY 80 VAL 81 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 ÍLE 79 VAL 81 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 1LE 79 GLY 80 VAL 81 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 SER 78? LE 79 VAL 81 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 ALA 74 LEU 75 ASN 76 ASN 77 BE 78? LE 79 GLY 80 LEU 82 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 ALA 73 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 ASN 77 ILE 79 OLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 ASN 77 SER 78 ELE 79 VAL 81 LEU 82 GLY 83 ALA 73 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 ALA 73 ASN 76 ASN 77 SER 78 ELE 79 GLY 80 VAL 81 GLY 83 ALA 73 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 77 ÍLE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 77 SER 78? LE 79 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 ALA 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 SER 78? LE 79 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 8! GLY 83 ALA 73 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 ÍLE 79 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77? LE 79 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ÍLE 79 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 - LE 79 GLY 80 LEU 82 WING 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 WING 73 WING 74 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 77 SER 78 - LE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 77 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 76 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 WING 74 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 WAT 76 WAT 78 WEST 79 WEST 80 W 81 WED 83 WED 73 WING 73 WED 74 WED 76 WED 78 WED 79 WEST 80 W 81 WED 82 WED 73 WED 74 WED 76 WED 77 WED 80 WED 81 WED 83 WED 83 W 73 ALA 74 ASN 76 ASN 77 ELE 79 VAL Yes LEU 82 GLY 83 ALA 73 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ASN 77? LE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 7 7 BE 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 WING 73 WING 74 ASN 76 ASN 77 SER 78 LE 79 GLY 80 VAL 81 ALA 73 ALA 74 LEU 75? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 1LE 79 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 1LE 79 GLY 80 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ÍLE 79 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ELE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ÍLE 79 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 76 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 7 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76? LE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ILE 79 GLY 80 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76? LE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 SER 78 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 SER 78 ILE 79 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 781LE 79 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 ÍLE 79 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 SER 78 ÍLE 79 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 76 ASN 77 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77? LE 79 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 ILE 79 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ELE 79 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ELE 79 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ELE 79 VAL 81 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 SER 78? LE 79 GLY 80 TABLE 10 NONUPLE mutation variants LEU 75 ASN 76 ASN 77 BE 78? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 ASN 76 ASN 77 BE 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 77 BE 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 SER 78? LE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 1LE 79 VAL 81 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 LE 79 GLY 80 LEU 82 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 BE 78 LE 79 GLY 80 VAL 81 GLY 83 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 BE 78 ANG 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 BE 78 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 LEU 75 ASN 76 ASN 77 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 WAND 76 WAND 78 1LE 79 GLY 80 WAND 81 LEU 82 GLY 83 WING 73 WING 74 WAND 76 WAND 78 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 ING 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 BE 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 AS N 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 SER 78 ELE 79 GLY 80 VAL 81 LEU 82 GLY 83 ALA 73 ALA 74 LEU 75 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78? LE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78? LE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ELE 79 WEL 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 GLY 80 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 BE 78 ÍLE 79 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 1LE 79 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 76 SER 78 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 GLY 80 VAL 81 LEU 82 GLY 83 WING 7 3 WING 74 LEU 75 ASN 76 ASN 77 WEA 79 W 81 W 82 W 83 W 73 W W 74 W 75 D 76 W 76 D 79 W 80 W 80 W 83 W 73 W 74 W 75 W 76 76,081 W 79 W 81 W 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 ILE 79 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 VAL 81 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 LEU 82 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 GLY 80 VAL 81 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 BE 78 ILE 79 LEU 82 GLY 83 ALA 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 IOL 81 GLY 83 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 VAL 81 LEU 82 ALA 73 ALA 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 GLY 83 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 LEU 82 WING 73 WING 74 LEU 75 ASN 76 ASN 77 SER 78 ILE 79 GLY 80 VAL 81 TABLE 11 7 One or more additional substitution mutations ("stabilizing substitutions") outside the epitope region can be further made to, for example, re-stabilize the protease during mutation of the epitope region or to increase the proteolytic activity of the variant. Many of these stabilizing mutations are well known in the art. Examples of stabilizing mutations are described, for example, in WO 95/10591, Baeck et al., Published April 20, 1995; patent of E.U.A. No. 4,914,031, Zukowski et al., Issued April 3, 1990; patent of E.U.A. No. 5,470,733, Brvan et al., Issued November 28, 1995; patent of E.U.A. No. 5,567,601, Brvan et al., Issued October 22, 1996; WO 89/07642, patent of E.U.A. No. 5,707,848, Brvan et al., Issued January 13, 1998; Van Eekelen et al., Published on August 24, 1989; WO 87/04461. Stabinskv et al., Published on July 30 and 1987; patent of E.U.A. No. 4,760,025, Estell et al., Issued July 26, 1988; WO 92/11348, Branner et al., Published July 9, 1992; EP 0,405,901, Casteleiin et al., Published January 2, 1991; WO 91/00345, Branner et al., Published January 10, 1991; and WO 94/10020, Brode et al., published March 23, 1995. Preferred stabilizer mutations include one or more of I107V; K213R; Y217L; Y217K; N218S; G169A; M50F; Q19E; P5A; S9A; 131 L; E156S; G169A; N212G; S188P; T254A; S3C + Q206C; and Q271E. More preferred stabilizing mutations include one or more of P5A; S9A; I31L; E156S; G169A; N212G; S188P; T254A; S3C + Q206C; Q271E; Y217L and Y217K. More preferred stabilizing mutations include Y217 and Y217K. A preferred variant for use in personal care compositions is? 75-83, Y217L.

Manufacturing Method Variants are prepared by mutation of the nucleotide sequences encoding the wild-type serine protease, thus resulting in variants having modified amino acid sequences. Such methods are well known in the art; one of these methods is as follows: A phagemid (pSS-5) containing the wild-type BNP 'subtilisin gene (Mitchison C. and JA Wells, "Protein Engineering of Disulfide Bonds in Subtilisin BPN"), Biochemistry, Vol. 28, pp. 4807-4815 (1989) is transformed into strain CJ236 of Escherichia coli dut-ung and a single DNA template containing single chain uracil is produced using the auxiliary phage of VCSM13 (Kunkel et al. ., "Rapid and Efficient Site-Specific Mutagenesis Without Phenotypic Selection", Methods in Enzymology, Vol 154, pp. 367-382 (1987), as modified by Yuckenbert et al., "Site-Directer in vitro Mutagenesis Using Uracil-Containing DNA and Phagemid Vectors, "Directed Mutagenesis - A Practical Approach, McPherson, MJ ed., Pp. 27-48 (1991). Site-directed mutagenesis modified from the method of Zoller and Smith (Zoller, MJ, and M Smith," Oligonucleotide-Directed Mutagenesis Using M13-Derived Vectors: An Efficient and General Procedure for the Production of Point Mutations in any DNA Fragment, "Nucleic acids Research, Vol. 10, pp. 6487-6500 (1982) is used to produce all mutants (essentially as presented by Yuckenberg et al., supra.) Oligonucleotides are made using a DNA synthesizer from 380B (Applied Biosystems Inc.). The mutagenesis reaction products are transformed into the Escherichia coli strain MM294 (American Type Culture Collection E. coli 33625). All mutations are confirmed by DNA sequencing and the isolated DNA is transformed into the expression strain PG632 of Bacillus subtilis (Saunders et al., "Optimization of the Signal-Sequience Cleavage Site for Secretion From Bacillus subtilis of a 34-amino acid. Fragment of Human Parathyroid Homone "; Gene, Vol. 102, pp. 277-282 (1991) and Yang et al.," Cloning of the neutral Protease Gene of Bacillus subtilis and the Use of the Cloned Gene to Créate an in w? ra-Derived Deletion Mutation ", Journal of Bacteriology, Vol. 160, pp. 15-21 (1984) .Preliminary evaluation of variant activity is determined by the ability of PG632 cells transformed with mutant plasmids to hydrolyze casein. does as follows: Cells Bacillus subtilis (PG632) containing the variant of interest are grown to mid-log phase in one liter of LB broth containing 10 g / L of glucose, and inoculated in a Biotat C fermentor (Braun Biotech, Inc., Allentown, PA) in a total volume of 9 liters. The fermentation medium contains yeast extract, casein hydrosylate, partially hydrolyzed starch, soluble (Maltrin M-250), anti-foam, pH regulators and trace minerals (see "Biology of Bacilli: Aplications to Industry", Doi ,. RH and M. McGloughlin, eds. (1992)). The broth is maintained at a constant pH of 7.5 during the fermentation operation. Kanamycin (50 μg / mL) is added for antibiotic selection of the mutagenized plasmid. The cells are cultured for 18 hours at 37 ° C at an Aβ0 of about 60 and the product is harvested. The fermentation broth is collected through the following steps to obtain a pure variant. Bacillus subtilis cells are removed from the broth by tangential flow against a membrane of 0.16 μm. The cellless broth is then concentrated by ultrafiltration with a molecular weight cut-off membrane of 8000. The pH is adjusted to 5.5 with concentrated MES pH regulator (2- (N-mofolino) etansu.phonic acid). The variant is then purified by cation exchange chromatography with S-sepharose and eluted with NaCl gradients. (See Scopes, RK, "Protein Purification Principles and Practice," Springer-Verlag, New York (1984) .A pNA test (DelMar et al., Analytical Biochemistry, Vol. 99, pp. 316-320 (1979)) is used to determine the concentration of active variant for fractions collected during gradient elution This test measures the rate at which p-nitroaniline is released as the variant hydrolyzes the soluble synthetic substrate, succinyl-alanine-alanine-proline phenylalanine-p-nitroaniline (sAAPF-pNA) The yellow-colored production rate of the hydrolysis reaction is measured at 410 nm in a spectrophotometer and is proportional to the concentration of active enzyme, In addition, absorbance measurements at 280 nm are used to determine the total protein concentration, the ratio of active enzyme / total protein gives the purity of the variant, and is used to identify fractions that have to be deposited for the supply solution. Utolysis of the variant during storage, weight of propylene glycol is added equal to the deposited fractions obtained from the chromatographic column. Upon completion of the purification procedure, the purity of the supply variant solution is verified with SDS-PAGE (sodium dodecylsulfate-polyacrylamide gel electrophoresis) and the absolute enzyme concentration is determined through a site titration method active using type II-T trypsin inhibitor: turkey egg white (Sima Chemical Company, St. Louis, Missouri). During preparation for use, the enzyme supply solution is eluted through an explosion-sized Sephadex-G25 column (Pharmaci, Piscataway, New Jersey) to remove the propylene glycol and exchange the pH regulator. The MES pH regulator in the enzyme supply solution is exchanged for the tris pH buffer of 0.1 M (tris (hydroxymethyl aminomethane) containing 0.01 M CaCl2 and the pH is adjusted to 8.6 with HCl. All experiments are carried out with pH of 8.6 in tris pH regulator with thermostat at 25 ° C.

Analytical Methods The variants of the present invention can be tested for enzymatic activity and immune and / or allergenic response using the following methods, which are known to those skilled in the art. Alternatively, other methods well known in the art can be used.

Activity of a variant The protease activity of a variant of the present invention can be tested by methods that are well known in the art. Such methods are discussed below: Skin flake activity method By using Scotch® tape # 3750G, human skin flakes are separated from the legs of a subject repeatedly until the tape is substantially opaque with flakes. The tape is then cut into squares of 2.54 by 2.54cm. and it is left aside. In a 10mm petri dish. by 35mm., 2mL of 0.75 mg / mL of a control enzyme (for example, subtilisin BPN ') or the variant to be tested is added the pH regulator of KH2PO4 0.01 M at a pH of 5.5. To this solution is added 1 mL of 2.5% sodium laurate at a pH of 8.6. The solution is mixed gently on a platform shaker. The box of previously prepared tape is soaked in the solution (the leaflet was up) for ten minutes continuing the smooth mixing. The tape box is then gently rinsed in tap water for fifteen seconds. Stevenel blue dye pipette (3 ml, commercially available from Sigma Chemical Co., St. Louis, MO) was applied in a clean petri dish. The rinsed tape box is placed in the dye for 3 minutes (with the leaflet facing up) with gentle mixing. The tape box is removed from the dye and rinsed consecutively in two 300 ml beakers with distilled water, for 15 seconds by rinsing. The tape box is allowed to air dry. The color intensity between the tape frame obtained from the control enzyme and the tape box obtained from the variant is visually compared or using a chromameter. In relation to the control enzyme tape frame, a variant tape box that shows less intensity of color indicates a variant that has higher activity.

Collagen-dyed activity method 50 ml of 0.1 M tris pH-regulator (tris-hydroxymethylaminomethane) containing 0.01 M CaCl2 are combined to give a pH of 86, and 0.5 grams of azocol (collagen impregnated with azo dye, commercially available from Sigma Chemical Co., St. Louis, MO). This mixture is incubated at 25 ° C while mixing gently with a platform shaker. 2 ml of the mixture is filtered through a 0.2 micron syringe cylinder and the absorbance of the mixture is read at 520 nm to zero a spectrophotometer. One ppm of a control enzyme (eg, subtilisin BPN ') of the variant to be tested is added to the remaining 48 ml of tris / azocol mixture. 2 ml of the control / variant containing solution is filtered through a syringe filter of 2 microns every two minutes for a total of ten minutes. For each filtered sample, the absorbance is read immediately at 520 nm. The results are plotted against time. The slopes of the control and the test conjugate indicate relative activities of the samples. A higher slope indicates a higher activity. The test variant activity (pending) can be expressed as percent of the control activity (pending).

T Cell Proliferation Test The immune and / or allergenic potential of variants of the variants of the present invention can be determined using a T cell proliferation test such as the test presented below. This test is a variation of the test described in Bungy Poor Fard et al .. "T Cell Epitopes of the Major Fraction of Rye Grass Lolium perennial (Lol p I) Defined Using Overlapping Peptides in vitro and in vivo", Clinical Experimental Immunology, Vol. 94, pp. 111-116 (1993). The blood of subjects allergic to subtilisin BPN '(positive to prick test) and control subjects (negative to prick test) are used in this test. The blood (~ 60 ml) of each subject is collected and the mononuclear cells are harvested using ficoll-hypaque (available from Pharmacia, Piscataway, New Jersey). Cells are washed twice in RPMI 1640 (obtainable from Gibco, Grand Island, New York) and then resuspended in complete RPMI medium supplemented with 10% 10% human AB serum, 2 mM L-glutamine. and 25 μg / mL of gentamicin (available from Gibco). The cells are cultured at a concentration of 2 x 10 5 cells / well in 0.2 ml of complete medium in 96-well microtitre plates with U-bottom. The potential antigen to be tested (either BPN 'inactivated as control positive or a variant of the present invention is added to a final concentration of up to 40 μg / ml Cultures are incubated at 37 ° C in 5% C02.After 5 days, 1 μCi / cavity of methyl-3H-thymidine The cells are harvested and later harvested.The incorporation of 3H-thymidine by the cells is evaluated as a measurement of T cell proliferation by liquid scintillation counting.

Compositions of the present invention Variants of the present invention can be used in cleaning compositions, including but not limited to laundry composition, hard surface cleaning compositions, light duty cleaning compositions including tableware cleaning compositions and detergent compositions. for automatic dishwashing. The cleaning compositions of the present invention comprise an effective amount of one or more variants of the present invention and a vehicle of cleaning composition. As used herein, "effective amount of variants", or the like, refers to the necessary amount of variant to achieve the necessary proteolytic activity in the specific cleaning composition. Said effective amounts are easily investigated by one of ordinary skill in the art and are based on many factors, such as the particular variant used, the cleaning application, the specific composition of the cleaning composition and if desired a liquid or dry composition (eg, granulate, stick), and the like. Preferably, the cleaning compositions are comprised of from about 0.0001% to about 10%, most preferably from about 0.001% to about 1% and, most preferably, from about 0.01% to about 0.1% of one or more variants of this invention . Several examples of different cleaning compositions in which the variants can be used are discussed in more detail below. In addition to these variants, these cleaning compositions further consist of a vehicle of the cleaning composition containing one or more materials of the cleaning composition compatible with the variant. The term 'cleaning composition material', as used herein, means any material selected for the particular type of cleaning composition desired and the shape of the product (eg, liquid, granulate, stick, aerosol, pellet). , in paste, in gel), whose materials are also compatible with the variant used in the composition. The specific selection of materials of the cleaning composition is carried out with ease considering the material to be cleaned and the desired form of the composition for cleaning conditions during use. The term 'compatible', as used herein, means that the materials of the cleaning composition do not reduce the proteolytic activity of the variant to such a degree that the variant is not as effective as desired during situations of normal use. The specific materials of the cleaning composition are exemplified in detail below. The variants of this invention can be used in a variety of detergent compositions in which it is desired to obtain a high degree of foaming and a good cleaning activity. Therefore, the variants can be used with several common ingredients in order to provide hard surface cleaners of complete formulation, dishwashing compositions, fabric washing compositions, and the like. Said compositions may be in liquid, granulated, bar and the like form. Such compositions can be formulated as 'concentrated' detergents containing from about 30% to about 60% by weight of surfactants. The cleaning compositions herein may optionally contain, and preferably, various surfactants (for example, anionic, nonionic or zwitterionic surfactants). Such surfactants are typically present at levels of about 5% to about 35% of the compositions. Non-limiting examples of the surfactants useful herein include the Cn-Ciß alkylbenzenesulfonates and the common primary and random alkyl sulfates, the secondary alkyl sulfates (2,3) of C? 0-C? 8 of the formulas CH3 (CH2)? (CHOSO3) "M +) CH3 and CH3 (CH2) and (CHOS? 3" M +) CH CH3, wherein x and (y + 1) are integers of at least about 7, preferably at least about 9, and M is a cation soluble in water, especially sodium; the alkylalkoxy sulfates of C? oC-? 8 (especially EO 1-5 ethoxysulfates), the alkylalkoxycarboxylates of C-C-is (especially the EO 1-5 ethoxycarboxylates), the alkyl polyglucosides of C-io-C-a and their corresponding sulphated polyglycosides, the α-sulfonated fatty acid esters of C 2 -C 8, the C 2 -C 2 alkyl and the alkylphenolalkoxylates (especially ethoxylates and ethoxy / mixed propoxy), the betaines and sulfobetaines sultaines') of C? 2-Ci8, the C10-C18 amine oxides, and the like. Preferred herein are alkylalkoxy sulfates (AES) and alkylalkoxycarboxylates (AEC). The use of such surfactants in combination with the amine oxide and / or betaine or sultaine surfactants is also preferred, depending on what the formulator desires. Other common useful surfactants are mentioned in normal texts. Particularly useful surfactants include the C 10 -C 18 N-methyl glucamides described in U.S. Pat. 5,194,639, Connor et al., Issued March 16, 1993. A wide variety of other ingredients useful in detergent cleansing compositions can be included in the compositions herein, including, for example, other active ingredients, carriers, hydrotropes, processing aids, dyes or pigments, and solvents for liquid formulations. If it is desired to obtain an additional foaming increase, they can be incorporated into the foam-enhancing compositions, such as C?-C? 6 alcoalcolamides, typically at levels of about 1% to about 10%. C? 0-C14 monoethanolamides and diethanolamides illustrate a typical class of such foam boosters. It is also convenient to use said foam enhancers with auxiliary foaming surfactants, such as the amine oxides, betaines and sultaines mentioned above. If desired, soluble magnesium salts, such as MgCl 2, MgSO 4 and the like, can be added, typically at levels of from about 0.1% to about 2% to provide additional foaming. The liquid detergent compositions herein may contain water and other solvents as carriers. The primary or secondary low molecular weight alcohols are exemplary, exemplified by methanol, ethanol, propanol and / so-propanol. Monohydric alcohols are preferred for solubilizing surfactants, but polyols, such as those containing from about 2 to about 6 carbon atoms and from about 2 to about 6 hydroxy groups (eg, 1,3-propanediol, ethylene glycol), can also be used. , glycerin and 1,2-propanediol). The compositions may contain from about 5% to about 90%, typically from about 10% to about 50%, of said vehicles. The detergent compositions herein will be formulated, preferably, such that during use in aqueous cleaning operations, the wash water contains a pH between about 6.8 and about 11. The finished products are generally formulated on this scale. Techniques for controlling pH at recommended levels of use include the use of, for example, pH, alkali and acid regulators. Such techniques are well known to those skilled in the art. In formulating the hard surface cleaning compositions and fabric cleaning compositions of this invention, the formulator may wish to employ various builders at levels of from about 5% to about 50% by weight. Typical builders include zeolites of 1 to 10 microns, polycarboxylates, such as citrates and oxydisuccinates, extralated silicates, phosphates and the like. Other common detergency builders are mentioned in the normal forms. Also, the formulator may wish to employ several additional enzymes, such as cellulases, lipases, amylases and proteases in said compositions, typically at levels from about 0.001% to about 1% by weight. Various detersive and fabric care enzymes are well known in the laundry detergent art. Various bleaching compounds, such as percarbonates, perborates and the like, can be used in such compositions, typically at levels of about 1% to about 15% by weight. If desired, said compositions may contain bleaching activators, such as tetraacetylethylenediamine, nonanoyloxybenzenesulfonate and the like, which are also known in the art. Usage levels vary, generally, from about 1% to about 10% by weight. Dirt releasing agents, especially of the anionic oligoester type; chelating agents, especially aminophosphonates and ethylenediamine disuccinates; clay soil removal agents, especially ethoxylated tetraethylenepentamine; dispersing agents, especially polyacrylates and polyaspartates; brighteners, especially anionic brighteners, suds suppressors, especially silicones and secondary alcohols; Fabric softeners, especially smectite clays, and the like, can be used in said compositions at levels ranging from about 1% to about 35% by weight. Normal forms and published patents contain multiple detailed descriptions of such common materials. Also, enzyme stabilizers can be used in the cleaning compositions. Said enzyme stabilizers include propylene glycol (preferably from about 1% to about 10%), sodium formate (preferably from about 0.1% to about 1%) and calcium formate (preferably from about 0.1% to about 1%). The variants of the present invention are useful in hard surface cleaning compositions. The expression 'hard surface cleaning compositions', as used herein, refers to liquid and granular detergent compositions for cleaning hard surfaces, floors, walls, bathroom tiles and the like. The hard surface cleaning compositions of this invention comprise an effective amount of one or more variants of this invention, preferably from about 0.001% to about 10%, most preferably from about 0.01% to about 5%, still more preferably about 0.05% to about 1% by weight variant composition. In addition to consisting of one or more variants, said hard surface cleaning compositions are generally constituted by surfactants and water soluble sequestering detergents. However, in certain specialized products, such as aerosol glass cleaners, the surfactants are sometimes not used, since they can produce a residue in the form of a layer and / or streaking on the surface of the glass. The surfactant component, when present, may comprise at least 0.1% of the compositions herein, but, generally, the compositions will contain from about 0.25% to about 10%, preferably from about 1% to about 5%. % surfactant. Commonly, the compositions will contain from about 0.5% to about 50% of a builder, preferably from about 1% to about 10%. Preferably, the pH should fall within the range of about 7 to about 12. Common pH adjusting agents, such as sodium hydroxide, sodium carbonate or hydrochloric acid, may be employed if an adjustment is required. Solvents may be included in the compositions. Useful solvents include, but are not limited to, glycol ethers, such as diethylene glycol monohexyl ether, diethylene glycol monobutyl ether, ethylene glycol monobutyl ether, ethylene glycol monohexyl ether, propylene glycol monobutyl ether, dipropylene glycol monobutyl ether, and diols, such as ethylene glycol. , 2,4-trimethyl-1,3-pentanediol and 2-ethyl-1,3-hexanediol. When used, said solvents are present, generally, at levels of from about 0.5% to about 15%, preferably from about 3% to about 11%. In addition, highly volatile solvents, such as / so-propanol or ethanol, can be used in these compositions to facilitate that the composition evaporates faster from the surfaces when the surface is not cleaned after a 'full concentration' application of the composition to the surface. When used, volatile solvents are usually present at levels of about 2% to about 12% in the compositions. The hard surface cleaning compositions of this invention are illustrated by the following examples.

EXAMPLES 1-6 Liquid compositions for hard surface cleaning All the formulas are adjusted to a pH of 7.

In Examples 1-6, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 75-82, with substantially similar results. In another embodiment of this invention, dishwashing compositions consist of one or more variants of this invention. The term 'dishwashing composition', as used herein, refers to, but is not limited to, all forms of dishwashing compositions, including granular and liquid forms. The dishwashing compositions of this invention are illustrated by the following examples.

EXAMPLES 7-10 Liquid dishwashing detergent All formulas are adjusted to a pH of 7. In Examples 7-10, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 79, with substantially similar results.

The liquid fabric cleaning compositions of this invention are illustrated by the following examples.

EXAMPLES 11-13 Liquid compositions for fabric cleaning In Examples 11-13, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 70, 75-82, with substantially similar results.

Compositions for personal care These variants are especially suitable for use in personal care compositions, such as rinse and rinse-off hair conditioners, shampoos, rinsing and non-rinsing anti-acne compositions, facial milks and conditioners, shower-bath gels, soaps , foaming and non-foaming facial cleansers, cosmetics, lotions and body, facial and hand moisturizers, non-rinsing facial moisturizers, cosmetic wipes and cleansers, oral care compositions and contact lens care compositions. These personal care compositions comprise one or more variants of this invention and a personal care vehicle. By way of illustration, these variants are suitable for inclusion in the compositions described in the following references: U.S. Patent No. 5,641, 479, Linares et al., Issued June 24, 1997 (cleansers for the skin); U.S. Patent No. 5,599,549 Wivell et al., Issued February 4, 1997 (cleansers for the skin); U.S. Patent No. 5,585,104, Ha et al., Issued December 17, 1996 (cleansers for the skin); U.S. Patent No. 5,540,852, Kefauver et al .. issued July 30, 1996 (cleansers for the skin); U.S. Patent No. 5,510,050, Dunbar et al., Issued April 23, 1996 (cleansers for the skin); U.S. Patent No. 5,612,324, Guang Lin et al., Issued March 18, 1997 (anti-acne preparations); U.S. Patent No. 5,587,176, Warren et al., Issued December 24, 1996 (anti-acne preparations); U.S. Patent No. 5,549,888, Venkateswaran. issued on August 27, 1996 (anti-acne preparations); U.S. Patent No. 5,470,884, Corless et al., Issued November 28, 1995 (anti-acne preparations); U.S. Patent No. 5,650,384, Gordon et al., Issued July 22, 1997 (gels for shower bath); U.S. Patent No. 5,607,678, Moore et al., Issued March 4, 1997 (gels for shower bath); U.S. Patent No. 5,624,666, Coffindaffer et al., Issued April 29, 1997 (conditioners and / or hair shampoos); U.S. Patent No. ,618,524, Bolich et al., Issued April 8, 1997 (conditioners and hair shampoos); U.S. Patent No. 5,612,301, Inman. Issued on March 18, 1997 (conditioners and hair shampoos); U.S. Patent No. 5, 573,709, Wells, issued November 12, 1996 (conditioners and / or hair shampoos); U.S. Patent No. 5,482,703, Pings, issued January 9, 1996 (conditioners and / or hair shampoos); U.S. Patent No. Re. 34, 584, Grote et al. Grote et al., Reissued on April 12, 1994 (conditioners and / or hair shampoos); U.S. Patent No. 5,641, 493, Date et al., Issued June 24, 1997 (cosmetics); U.S. Patent No. 5,605,894, Blank et al., Issued February 25, 1997 (cosmetics); U.S. Patent No. 5,585,090, Yoshioka et al., Issued December 17, 1996 (cosmetics); U.S. Patent No. 4,939,179, Chenev et al., Issued July 3, 1990 (lotions for hands, face and / or body); U.S. Patent No. 5,607,980, McAtee et al., Issued March 4, 1997 (lotions for hands, face and / or body); U.S. Patent No. 4,045,364, Richter et al., Issued August 30, 1997 (cosmetic cloths and cleansers); European patent application EP 0619074, Touchet et al., published on October 12, 1994 (cosmetic cloths and cleansers); U.S. Patent No. 4,475,217, Brown-Skrobot et al., Issued December 4, 1990 (cosmetic cloths and cleansers); U.S. Patent No. 5,096,700, Seibel. Issued on March 17, 1992 (compositions for oral cleaning); U.S. Patent No. 5,028,414, Sampathkumar, issued July 2, 1991 (compositions for oral cleaning); U.S. Patent No. 5,028,415, Benedict et al., Issued July 2, 1991 (compositions for oral cleaning); U.S. Patent No. 5,028,415, Benedict et al., Issued July 2, 1991 (compositions for oral cleaning); U.S. Patent No. 4,863,627, Davies et al .. September 5, 1989 (cleaning solutions for contact lenses); U.S. Patent No. Re. 32,672, Huth et al., Reissued on May 24, 1988 (cleaning solutions for contact lenses); and U.S. Patent No. 4,609,493, Schafer, issued September 2, 1986 (cleaning solutions for contact lenses). To further illustrate the oral cleaning compositions of this invention, an amount of one or more pharmaceutically acceptable variants of this invention is included in compositions useful for removing proteinaceous stains from teeth or dentures. The term 'compositions oral cleansing', as used herein, refers to dentifrices, toothpastes, tooth gels, tooth powders, mouthwashes, mouth sprays, mouth gels, chewing gums, tablets, pouches, tablets, biogels, prophylactic pastes, dental treatment solutions and the like. Preferably, the oral cleaning compositions consist of from about 0.0001% to about 20% of one or more variants of this invention, most preferably from about 0.001% to about 10%, most preferably still from about 0.01% to about 5% in composition weight, and a pharmaceutically acceptable vehicle. The term "pharmaceutically acceptable", as used herein, means that the drugs, medicaments or inert ingredients described by the term are suitable for use in contact with human and lower animal tissues without risking undue toxicity, incompatibility. , instability, irritation, allergic response and the like, in proportion to a reasonable benefit / risk ratio. Typically, the pharmaceutically acceptable components of the buccal cleaning vehicle of the buccal cleaning components of the buccal cleaning compositions will generally be comprised of from about 50% to about 99.99%, preferably from about 65% to about 99.99%, most preferably from about 65% to about 99%, by weight of the composition. The pharmaceutically acceptable components of the vehicle and the optional components that can be included in the buccal cleaning compositions of this invention are well known to those skilled in the art. A wide variety of types of compositions, vehicle components and optional components useful in oral cleaning compositions is described in the references cited above. In another embodiment of this invention, denture cleansing compositions for cleaning dentures outside the buccal cavity comprise one or more variants of this invention. Said denture cleansing compositions consist of an effective amount of one or more variants, preferably from about 0.0001% to about 50% of one or more variants, most preferably from about 0.001% to about 35%, even more preferably about 0.01. % to about 20%, by weight of the composition, and a denture cleansing vehicle. Various formats of denture cleansing compositions, such as effervescent tablets and the like, are well known in the art (see, for example, U.S. Patent No. 5,055,305, Young) and, in general, are suitable for incorporating one or more variants for Remove proteinaceous spots from dentures. In another embodiment of this invention, contact lens cleaning compositions consist of one or more variants of this invention. Said contact lens cleaning compositions contain an effective amount of one or more variants, preferably from about 0.01% to about 50% of one or more variants, most preferably from about 0.01% to about 20%, even more preferably from about 1% to about 5%, by weight of the composition, and a contact lens cleaner vehicle. Various formats of contact lens cleaning compositions, such as tablets, liquids and the like, are well known in the art and are generally suitable for incorporating one or more variants of this invention to remove proteinaceous stains from contact lenses.

The contact lens cleaning composition embodiment of this invention is illustrated by examples 14-17.

EXAMPLES 14-17 Contact lens cleaning solution In Examples 14-17, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 75-82, with substantially similar results. Examples 18-21 illustrate the use of these variants in products for washing the body: EXAMPLES 18-21 Products for washing the body In Examples 18-21, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 75-83, Y217L, with substantially similar results. Examples 18-21 illustrate the use of these variants in facial cleansers: EXAMPLES 22 TO 25 Products for washing the face In Examples 22-25, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 73, I79M, with substantially similar results.

Examples 26-27 illustrate the use of these variants in non-rinsing skin-moisturizing compositions: EXAMPLES 26-27 Non-wettable composition of skin moisturizer In Examples 26-27, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 75-83, with substantially similar results. Example 28 illustrates the use of these variants in compositions applied to cleaning cloths.

EXAMPLE 28 Composition in cleaning cloths The above composition is impregnated in an absorbent woven cloth composed of cellulose and / or polyester at about 250% by weight of the absorbent cloth. In Example 28, the variants cited in Tables 2-10, and preferred variants cited herein, among others, replace? 75-82, G70V, with substantially similar results.

LIST OF SEQUENCES (1. GENERAL INFORMATION: (i) APPLICANT: RUBINGH, D.N. et al. (ii) TITLE OF THE INVENTION: SERINE PROTEASE VARIANTS THAT HAVE DECREASED IMMUNOGENICITY (iii) NUMBER OF SEQUENCES: 1 (iv) ADDRESS OF CORRESPONDENCE: (A) RECIPIENT: THE PROCTER & GAMBLE COMPANY (B) STREET: 11810 EAST MIAMI RIVER ROAD (C) CITY: ROSS (D) STATE: OH (E) COUNTRY: E.U.A. (F) POSTAL CODE: 45061 (v) COMPUTER LEADABLE FORM: (A): TYPE OF MEDIUM: Flexible disk (B) COMPUTER: IBM compatible PC (C) OPERATING SYSTEM: PC-DOS / MS-DOS (D) SOFTWARE: Patentln Relay # 1.0, Version # 1.30 (vi) CURRENT DATA OF THE APPLICATION: (A) APPLICATION NUMBER: (B) DATE OF SUBMISSION: (C) CLASSIFICATION: (viii) INFORMATION OF THE APPORTER / AGENT: (A) NAME: HERSKO, BART S. (B) REGISTRATION NUMBER: 32,572 (C) CASE NUMBER: (ix) TELECOMMUNICATIONS INFORMATION: (A) TELEPHONE: 513-627-0633 (B) TELEFAX: 513-627-0260 (2) INFORMATION FOR SEQ ID NO: 1: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 275 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 1: Ala Gln Ser Val Pro Tyr Gly Val Ser Gln He Lys Ala Pro Ala Leu 1 5 10 15 His Ser Gln Gly Tyr Thr Gly Ser Asn Val Lys Val Wing Val He Asp 20 25 30 Ser Gly He Asp Ser Ser His Pro Asp Leu Lys Val Wing Gly Gly Wing 35 40 45 Ser Met Val Pro Ser Glu Thr Asn Pro Phe Gln Asp Asn Asn Ser His 50 55 60 Gly Thr His Val Ala Gly Thr Val Ala Ala Leu Asn Asn Ser He Gly 65 70 75 80 Val Leu Gly Val Ala Pro Ser Ala Ser Leu Tyr Ala Val Lys Val Leu 85 90 95 Gly Wing Asp Gly Ser Gly Gln Tyr Being Trp He He Asn Gly He Glu 100 105 110 Trp Wing Wing Asn Asn Met Asp Val He A = n Met Ser Leu Gly Gly 115 120 125 Pro Ser Gly Ser Ala Ala Leu Lys Ala Ala Val Asp Lys Wing Val Wing 130 135 140 Ser Gly Val Val Val Val Wing Wing Wing Gly Asn Glu Gly Thr Ser Gly 145 150 155 160 Being Ser Thr Val Gly Tyr Pro Gly Lys Tyr Pro Ser Val He Wing 165 170 175 Val Gly Ala Val A = p Ser Ser Asn Gln Arg Wing Ser Phe Ser Ser Val 180 185 190 Gly Pro Glu Leu Asp Val Met Wing Pro Gly Val Ser He Gln Ser Thr 195 200 205 Leu Pro Gly Asn Lys Tyr Gly Wing Tyr Asn Gly Thr Ser Met Ala Ser 210 215 220 Pro His Val Ala Gly Ala Ala Ala Leu He Leu Ser Lys His Pro Asn 225 230 235 240 Trp Thr Asn Thr Gln Val Arg Ser Ser Leu Glu A = n Thr Thr Thr Lys 245 250 255 Leu Gly Asp Ser Phe Tyr Tyr Gly Lys Gly Leu He Asn Val Gln Wing 260 265 270 Wing Wing Gln 275

Claims

NOVELTY OF THE INVENTION CLAIMS

1. - A variant of a serine protease characterized by a modified amino acid sequence of a wild-type amino acid sequence, wherein the modified amino acid sequence comprises a deletion of one or more of positions 70-84 corresponding to subtilisin BPN ' , in which the variant is not? 75-83 or? 71.

2. A variant according to claim 1, further characterized in that the serine protease is selected from the group consisting of subtilisin BPN ', subtilisin Carisberg, subtilisin DY, subtilisin 309, proteinase K and termitase.

3. A variant according to claim 1 or 2, further characterized in that the deletion is one or more of positions 73-83 corresponding to subtilisin BPN '.

4. A variant according to any of the preceding claims, further characterized in that the deletion is one or more of positions 70 and 75-82, corresponding to subtilisin BPN '.

5. A variant according to any of the preceding claims selected from the group consisting of: (a)? 70.75-82; (b)? 75-82; (c)? 70.78.79; (d)? 70; (e)? 75; (f)? 76; (g)? 78; (h)? 79; (i)? 81; and G)? 82

6. - A variant according to any of the preceding claims, further characterized in that it consists of a substitution of one or more of positions 70-84, corresponding to subtilisin BPN '.

7. A variant according to any of the preceding claims, further characterized by consisting of one or more stabilizing mutations.

8. A mutant serine protease gene encoding a variant according to any of the preceding claims.

9. A personal care composition characterized by: (a) a variant of a serine protease having a modified amino acid sequence of a wild-type amino acid sequence, further characterized in that the modified amino acid sequence is constituted by a deletion of one or more of positions 70-84, corresponding to BPN '; and (b) a vehicle of the personal care composition.

10. A composition according to claim 9, wherein the variant is further characterized by a substitution in one or more of positions 70-84, corresponding to subtilisin BPN '.