KR890014743A - New polypeptide having growth factor activity and amino acid sequence encoding the polypeptide - Google Patents

Abstract

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Description

New polypeptide having growth factor activity and amino acid sequence encoding the polypeptide

Since this is an open matter, no full text was included.

1 shows a comparison of known amino acid sequence of growth factors and vaccinia virus. The first amino acid of VGF is aspartic acid (D) at position 20 from methionine corresponding to aa ^-24 of the published peptide.

2 shows the limited structure of mature VGF to which asparagine (N) will be replaced. Glycosylation potential sites of MGF and SFGF are shown with a star (*).

Claims (37)

(1) bind to the EGF receptor, (2) have substantially the same ring structure as the naturally occurring growth factor (“natural ligand”) capable of binding to the EGF receptor, and (3) each region is the same or mutually Having at least about 30% identity with a fragment of another naturally occurring ligand and having at least about 30% identity with a native ligand in a polypeptide consisting of three regions defined as follows: an amino acid sequence having the same relative position as the fragment of said naturally occurring ligand A DNA sequence encoding a polypeptide, wherein the three regions are the ^first region of at least about 11 central fragment amino acids starting at an amino acid near aa ^-6 -aa ¹ and ending at an amino acid near aa ¹¹ -aa ¹⁵ , aa ¹¹ -aa ¹⁵ starting at the amino acid aa in the vicinity of and at least about 11 up to near the end of a ²⁵ -aa ²⁹ of the central fragment amino A second region, starting at the amino acid aa ²⁵ -AA ²⁹ to the vicinity of the aa ⁴⁰ -aa ⁵³ is at least about 14 amino C- end of the third zone to the short amino acid which terminates in the vicinity, and wherein the DNA sequence is aa ^-7 - a fourth region of at least about six maximal N-terminal amino acids starting near aa ^-1 and ending near the amino acids aa ^-45 -aa ^-7 is optionally inserted into the carboxy terminal group of the fourth region and the polypeptide is When having the same amino acid sequence as the natural ligand, it is selectively separated with the amino terminal group of the first region assuming that there is a DNA sequence encoding the fourth region and that it encodes something different from the maximum N-terminal amino acid of the natural ligand. It is optionally further coded as possible amino acid sequences. The DNA sequence of claim 1, wherein the natural ligand is VGF, TGF-α, MGF, EGF, SFGF, or amphiregulin. The amino terminal group of the fourth region according to claim 1, wherein the DNA sequence inserts a leader sequence, optionally into a carboxy terminal group of the leader sequence, and when the amino terminal group of the first region or the fourth region is present DNA sequence, characterized in that it is further encoded as a selectively separable amino acid sequence bound to. 4. The DNA sequence of claim 3, wherein the leader sequence is a λ N-protein, a λ cro protein, or an N-terminal fragment of Ampiregulin. 4. The DNA sequence of claim 3, wherein the leader sequence is a signal sequence or variant thereof. 6. The DNA sequence of claim 5, wherein the signal sequence is an alkaline phosphatase signal sequence, a TGF-β signal sequence in monkeys, or a K. lactis spore toxin signal sequence. 8. The DNA sequence of claim 1, wherein at least one of the regions is at least about 30% identical to a fragment of VGF, TGF-β, EGF or SFGF. 9. 7. The DNA sequence of any one of claims 1 to 4, wherein said DNA sequence encodes VGF. 8. The DNA sequence of any one of claims 1, 4 and 7, wherein said DNA sequence encodes a TGP. 7. The DNA sequence of any one of claims 1, 4 and 6, wherein said DNA sequence encodes EGF. 2. The DNA sequence of claim 1, wherein said DNA sequence encodes SFGF. The DNA sequence of claim 1, wherein the sequence has at least one modification from the sequence encoding the naturally-occurring ligand changed as follows, wherein the HGD is HGT and GMYC is GYAC. RCS is replaced by VCS, CLHGDC is replaced by CLHGTC, and GMYCRC is replaced by GYACVC. From a bacteriophage λ N-protein or cro protein, ampyregulin, T4 gene 32, or bacterial alkaline phosphatase signal sequence, a sequence of fragments of naturally occurring growth factors that each region can bind to the EGF receptor (“natural ligand”) Conjugated via, via an optionally separable amino acid sequence, an amino terminal group of a polypeptide of interest consisting of three regions substantially defined as follows and having the same relative positions as the fragment of the naturally occurring ligand: in, including the variant of about 8-45 amino acid N- terminal groups of the polypeptide and a DNA sequence encoding the leader arrangement, starting in the vicinity of the three regions of the amino acid aa ^-6 -aa ¹ and amino acid aa ¹¹ - the first, region, amino acid aa ¹¹ -aa ¹⁵ that is at least about 11 N-terminal amino acids terminating near aa ¹⁵ Starting at the root, and the amino acid aa ²⁵ -aa ²⁹ of at least about 11 amino acid fragment of a second intermediate region terminating at about amino acid aa ²⁵ -aa ²⁹ -aa ⁴⁷ start at about ⁴⁰ and aa, at least about 14 amino terminating in the vicinity of And a third region, which is a C-terminal amino acid. 14. The DNA sequence of claim 13, wherein said leader sequence is about 32 or 33 N-terminal amino acids of said N-gene. The DNA sequence of claim 14, wherein said polypeptide of interest is TGF-α, VGF, VGFA, VGFa or EGF. 14. The DNA sequence of claim 13, wherein the leader sequence is an alkaline phosphatase signal sequence or a modified alkaline phosphatase signal sequence. 17. The DNA sequence of claim 16, wherein said polypeptide of interest is VGFa, nVGFa, or EGF. 15. The DNA sequence of claim 14, wherein said polypeptide of interest has the following amino acid sequence, wherein said amino acid sequence is M V V S H F N D C P D S H T Q F C F H G T C R F L V Q E E K P A C V C S H G Y T G I R C Q H V V L V D Y Q R 15. The DNA sequence of claim 14, wherein said polypeptide of interest has the following amino acid sequence, wherein said amino acid sequence is M D I P A I L L C G P E G D G Y C L H G T C R F L V Q E E P A C Y C S H G Y T G I R C Q H W L V D Y Q R 15. The DNA sequence of claim 14, wherein said polypeptide has the following amino acid sequence, wherein said amino acid sequence is M V V S H F N D C P D S H T Q F C F H G T C I H A R D I D G Y A C V C S H G Y T G I R C Q H W L V D Y Q R The DNA sequence of claim 13, wherein said leader sequence is about 21 N-terminal amino acids of said Cro protein. 20. The DNA sequence of claim 19, wherein the polypeptide of interest has the following amino acid sequence, wherein the sequence in the amino acid sequence is M V V S H F N D C P D S H T Q F C F H G T C R F L V Q E E K P A C V C S H G Y T G I R C Q H V V L V D Y Q R 14. The DNA sequence of claim 13, wherein said leader sequence is about 8 N-terminal amino acids of T4 gene 32. 24. The DNA sequence of claim 23, wherein said polypeptide of interest has the following amino acid sequence, wherein said amino acid sequence is V V S H F N D C P D S H T Q F C F H G T C R F L V Q E E K P A C V C H G Y T G I R C Q H V V L V D Y Q R. In the remnant direction, functional transcriptional and translational initiation regulatory regions in the host cell and (1) naturally occurring growth factors capable of binding to EGF receptors and (2) binding to EGF receptors (“natural ligands”) And (3) an amizoic acid array having substantially the same ring structure, and (3) each region having at least about 30% identity with the same or different fragments of the native ligand and having the same relative position as the fragment of the native ligand An expression cassette consisting of a DNA sequence encoding a polypeptide characterized by having at least about 30% identity with a naturally occurring ligand in a polypeptide consisting of three regions defined above, wherein the three regions are amino acids in the vicinity of aa- ^6- aa ¹ The first region of at least about 11 central fragment amino acids starting at and ending at amino acids near aa ¹¹ -aa ¹⁵ , a second region of at least about 11 central fragment amino acids starting at an amino acid near aa ¹¹ -aa ¹⁵ and ending at aa ²⁵ -aa ²⁹ , starting at an amino acid near aa ²⁵ -AA ²⁹ and aa ⁴⁰ -aa ⁵³ Terminating nearby, or the third region of at least about 14 amino C-terminal amino acids, and the DNA sequence starts around aa- ^7- aa ^{- 1} and terminates near amino acids aa- ^45- aa- ⁷ A DNA sequence in which the fourth region of at least about 6 maximum N-terminal amino acids is optionally inserted into the carboxy terminal group of the fourth region and the fourth region is loaded when the polypeptide has the same amino acid sequence as the natural ligand Is assumed to encode a functional transcriptional and detoxification regulatory region in the host cell different from the maximal N-terminal amino acid of the native ligand and the host cell. Optionally further encoding by selectively separable amino acid sequence in a state of being bonded to the amino end groups of the first region. In the direction of residue, a naturally occurring growth factor capable of binding functional transcriptional and translational initiation reactions in the host cell and (1) binding to EGF receptors, and (2) binding to EGF receptors (“natural ligands”). (3) an amino acid sequence having substantially the same ring structure, and (3) each region is at least about 30% identical to a fragment of the same or different naturally occurring ligand and has the same relative position as the naturally occurring ligand and the fragment A host cell consisting of an expression cassette comprising a DNA sequence encoding a polypeptide, characterized in that it has at least about 30% identity with a naturally occurring ligand in a polypeptide consisting of three regions defined above, wherein the three regions are aa- ^6- aa ^1. starting at the amino acid in the vicinity of and aa ¹¹ -aa ¹⁵ near at least about 11 central fragment terminating at amino acids A first region with amino acid, aa ¹¹ -aa starting at amino acid ¹⁵ near the start and at least a second zone at about 11 central fragment amino acids, amino acid aa ²⁵ -AA ²⁹ near the end in the vicinity of aa ²⁵ -aa ²⁹ and a third region of at least about 14 amino C-terminal amino acids terminating around aa ⁴⁰ -aa ⁵³ , and the DNA sequence starts around aa ^-7 -aa ^-1 and is of amino acid aa ^-40 -aa ^-7 A fourth region of at least about six maximal N-terminal amino acids terminating in the vicinity, optionally inserted into the carboxy terminal group of the fourth region, encoding the fourth region when the polypeptide has the same amino acid sequence as the native ligand The amino acid terminal of the first region is assuming that the DNA sequence is present and loads something different from the local maximum N-terminal amino acid Optionally further encoding by selectively separable amino acid sequence as hapdoen state. (1) have a ring structure substantially the same as a naturally occurring growth factor ("natural ligand") capable of binding to EGF receptors, (2) binding to EGF receptors, and (3) each region is the same or different At least about 30% identity to a native EGF receptor conjugate in a polypeptide consisting of three regions, defined as follows, having at least about 30% identity with a fragment of the native ligand and an amino acid sequence having the same relative position as the fragment of the native ligand A method for producing a polypeptide comprising: wherein the three regions begin with an amino acid near aa- ^6- aa ¹ and end with an amino acid near aa ¹¹ -aa ¹⁵ . A first region of at least about 11 central fragment amino acids, a second region of at least about 11 central fragment amino acids starting at an amino acid near aa ¹¹ -aa ¹⁵ and ending at aa ²⁵ -aa ²⁹ , aa ^25- a third region consisting of at least about 14 amino C-terminal amino acids starting at an amino acid near aa ²⁹ and ending at aa ⁴⁰ -aa ⁵³ and optionally inserted into the carboxy terminal group of the fourth region, the polypeptide Has the same amino acid sequence as the natural ligand, the first region is present and is different from the maximal N-terminal amino acid of the natural ligand and under the assumption that it is a functional transcriptional and translational terminal regulatory site in the host cell. start near aa ^-7 -aa ^-1 with optional detachable coupling to the amino acid sequence as the amino end groups of the zone status And amino acid aa _-45 -aa consists of a fourth region of the short-term amino acids is at least about six maximally N- end terminating in the vicinity of _7, in the above method, the method, suitable culture times that expressed poly-lactide the rep From the sustained to the transcriptional direction, a host cell comprising a functional transcriptional and translational initiation reaction regulatory region in the host cell, a DNA sequence tilated the polypeptide, and an expression cassette containing an electron and translation termination regulation region. And growing the polypeptide and isolating the polypeptide. pBMII / NDP / EGP, pBMII / PAD / EGF, pBMII / PAK / EGF, pBMII / N / TGF, pBMII / NP / VGFA, pBMII / NDP / VGFa, pBMII / PAD / nVGFa, pBMII / PAK / nVGFa, pRSV / VGF, pAc / SFGF, pAc / VGF, ptac / TTV, TacPak / EGF, pTCPt / EGF, pTCPt / nVGFa, pBMII / N / TTV, pBMII / N / TTV, pMII / NDP / TTV, pBMII / DNP / VTV, Or plasmid consisting of the functional regions of pVMI6 / TVV. (1) bind to the EGF receptor, (2) have substantially the same ring structure as the naturally occurring growth factor (“natural ligand”) capable of binding to the EGF receptor, and (3) each region is the same or mutually At least about 30% identity to a native EGF receptor conjugate in a polypeptide consisting of three regions defined as follows having an amino acid sequence having at least about 30% identity with a fragment of another native ligand and having the same relative position as the fragment of the native ligand A polypeptide comprising: a ^first of at least about 11 central fragment amino acids starting at an amino acid near the native aa- ^6- aa ¹ in the polypeptide comprising the three regions and ending at an amino acid near aa ¹¹ -aa ¹⁵ second region, starting at the amino acid aa ¹¹ -aa ¹⁵ near and at least terminating in the vicinity of aa ²⁵ -aa ²⁹ 11 central fragment amino acids of the second area, starting at the amino acid aa ²⁵ -aa ²⁹ -aa ⁵³ near to aa ⁴⁰ is at least about 14 amino C- end of the third area as a short-term amino acids terminating in the vicinity of, and optionally Is inserted into the carboxy terminal group of the fourth region, when the polypeptide has the same amino acid sequence as the natural ligand, the fourth region is present and different from the maximal N-terminal amino acid of the natural ligand and in the host cell. Under the assumption that it is a functional transcriptional and translational terminal regulatory region, the amino acid aa ^-45 -aa ^-7 is started in the vicinity of aa ^-7 -aa ^-1 having a selectively separable amino acid sequence bound to the amino acid terminal group of the first region. Consisting of a fourth region of at least about 6 maximum N-terminal amino acids ending in the vicinity of. A polypeptide consisting of a fragment of at least 37 amino acids consisting of at least about 1-37 amino acids or an amino acid sequence X ¹ is one bond or amino acid, X ² is one bond or 1-2 amino acids, X ³ is one bond or 1-4 amino acids, and X ¹ , X ² and X ³ are the same or mutually May vary. ^31. The polypeptide of claim 30, wherein at least one of aa ¹⁴ is threonine, the aa ²⁴ is tyrosine, aa ²⁵ is alanine and the aa ²⁷ is valine. 31. The polypeptide of claim 30, wherein said amino acid sequence consists of aa- ^6- aa ⁴⁷ . 33. The polypeptide of claim 32, wherein said aa ¹⁴ is threonine, said aa ²⁴ is tyrosine and said aa ²⁵ is alanine. 34. A polypeptide according to claim 33, wherein in the amino acid sequence, aa- ^6- aa ¹³ of the amino acid sequence is replaced as follows: being. 34. The polypeptide of claim 33 wherein aa ¹⁴ -aa ²⁷ of the amino acid sequence is replaced as follows: being. The polypeptide of claim 35, wherein aa ²² is replaced with glutamic acid. A DNA sequence encoding a polypeptide having any one of claims 30 to 36. ※ Note: The disclosure is based on the initial application.