CN101641433A - The cleaning compositions that contains transglucosidase - Google Patents

The cleaning compositions that contains transglucosidase Download PDF

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Publication number
CN101641433A
CN101641433A CN200880009337A CN200880009337A CN101641433A CN 101641433 A CN101641433 A CN 101641433A CN 200880009337 A CN200880009337 A CN 200880009337A CN 200880009337 A CN200880009337 A CN 200880009337A CN 101641433 A CN101641433 A CN 101641433A
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Prior art keywords
transglucosidase
composition
natural gum
cleaning
gum polysaccharide
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CN200880009337A
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Inventor
H·C·麦克唐纳
A·J·保罗斯
J·K·舍蒂
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Danisco USA Inc
Danisco US Inc
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Danisco USA Inc
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    • CCHEMISTRY; METALLURGY
    • C11ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11DDETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
    • C11D3/00Other compounding ingredients of detergent compositions covered in group C11D1/00
    • C11D3/16Organic compounds
    • C11D3/20Organic compounds containing oxygen
    • C11D3/22Carbohydrates or derivatives thereof
    • CCHEMISTRY; METALLURGY
    • C11ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11DDETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
    • C11D3/00Other compounding ingredients of detergent compositions covered in group C11D1/00
    • C11D3/16Organic compounds
    • C11D3/38Products with no well-defined composition, e.g. natural products
    • C11D3/386Preparations containing enzymes, e.g. protease or amylase
    • C11D3/38636Preparations containing enzymes, e.g. protease or amylase containing enzymes other than protease, amylase, lipase, cellulase, oxidase or reductase
    • CCHEMISTRY; METALLURGY
    • C11ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11DDETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
    • C11D3/00Other compounding ingredients of detergent compositions covered in group C11D1/00
    • C11D3/16Organic compounds
    • C11D3/20Organic compounds containing oxygen
    • C11D3/22Carbohydrates or derivatives thereof
    • C11D3/222Natural or synthetic polysaccharides, e.g. cellulose, starch, gum, alginic acid or cyclodextrin
    • CCHEMISTRY; METALLURGY
    • C11ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11DDETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
    • C11D3/00Other compounding ingredients of detergent compositions covered in group C11D1/00
    • C11D3/16Organic compounds
    • C11D3/38Products with no well-defined composition, e.g. natural products
    • C11D3/386Preparations containing enzymes, e.g. protease or amylase
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/10Transferases (2.)
    • C12N9/1048Glycosyltransferases (2.4)
    • C12N9/1051Hexosyltransferases (2.4.1)
    • C12N9/1071,4-Alpha-glucan branching enzyme (2.4.1.18)
    • DTEXTILES; PAPER
    • D06TREATMENT OF TEXTILES OR THE LIKE; LAUNDERING; FLEXIBLE MATERIALS NOT OTHERWISE PROVIDED FOR
    • D06LDRY-CLEANING, WASHING OR BLEACHING FIBRES, FILAMENTS, THREADS, YARNS, FABRICS, FEATHERS OR MADE-UP FIBROUS GOODS; BLEACHING LEATHER OR FURS
    • D06L1/00Dry-cleaning or washing fibres, filaments, threads, yarns, fabrics, feathers or made-up fibrous goods
    • D06L1/12Dry-cleaning or washing fibres, filaments, threads, yarns, fabrics, feathers or made-up fibrous goods using aqueous solvents
    • D06L1/14De-sizing
    • DTEXTILES; PAPER
    • D06TREATMENT OF TEXTILES OR THE LIKE; LAUNDERING; FLEXIBLE MATERIALS NOT OTHERWISE PROVIDED FOR
    • D06LDRY-CLEANING, WASHING OR BLEACHING FIBRES, FILAMENTS, THREADS, YARNS, FABRICS, FEATHERS OR MADE-UP FIBROUS GOODS; BLEACHING LEATHER OR FURS
    • D06L4/00Bleaching fibres, filaments, threads, yarns, fabrics, feathers or made-up fibrous goods; Bleaching leather or furs
    • D06L4/40Bleaching fibres, filaments, threads, yarns, fabrics, feathers or made-up fibrous goods; Bleaching leather or furs using enzymes

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Molecular Biology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Genetics & Genomics (AREA)
  • Zoology (AREA)
  • Textile Engineering (AREA)
  • Emergency Medicine (AREA)
  • Microbiology (AREA)
  • Biotechnology (AREA)
  • Biomedical Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Detergent Compositions (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Cleaning By Liquid Or Steam (AREA)

Abstract

The invention provides the composition that comprises transglucosidase and natural gum polysaccharide, wherein the natural gum polysaccharide is the substrate of this transglucosidase.The present invention also provides the method with transglucosidase degraded natural gum polysaccharide.In some preferred embodiments, find that said composition and method can be used in the cleaning applications.

Description

The cleaning compositions that contains transglucosidase
Invention field
[01] the invention provides the composition that comprises transglucosidase and natural gum polysaccharide, wherein said natural gum polysaccharide is the substrate of described transglucosidase.The present invention also provides the method for using transglucosidase degraded natural gum polysaccharide.In some preferred embodiments, find that said composition and method can be used in the cleaning applications.
Background of invention
[02] washing composition and other cleaning compositions often comprise the complex combination of effective constituent.For example, many cleaning product contain surfactant system, cleaning enzymes, SYNTHETIC OPTICAL WHITNER, washing assistant, suds suppressor, soil-suspending agent, stain remover, white dyes, softening agent, dispersion agent, dye transfer inhibition compound, abrasive, sterilant and spices.Yet,, have a lot of spots still to be difficult to remove although current washing composition is so complicated.Therefore, still exist in this area the composition of the various spots of effective cleaning and the demand of method.
Summary of the invention
[03] the invention provides the composition that comprises transglucosidase and natural gum polysaccharide, wherein, this natural gum polysaccharide is the substrate of this transglucosidase.The present invention also provides the method for using transglucosidase degraded natural gum polysaccharide.In some preferred embodiments, found said composition and the method purposes in cleaning applications.
[04] in some embodiments, above-mentioned composition comprises the natural gum polysaccharide such as xanthan gum etc.In some further embodiments, transglucosidase has the activity that is defined as EC 2.4.1.24 according to the IUBMB nomenclature.In some other embodiment, transglucosidase has and the aminoacid sequence of Aspergillus (Aspergillus) the transglucosidase aminoacid sequence at least about 90% identity.Further in the embodiment, composition also comprises at least a tensio-active agent at some.In some other embodiment, the natural gum polysaccharide exists as the spot on the object.In some particularly preferred embodiments, object is a fabric.
[05] the present invention also provides and has comprised transglucosidase is combined method with degraded natural gum polysaccharide with at least a natural gum polysaccharide.In some embodiments, transglucosidase has the activity that is defined as EC 2.4.1.24 according to the IUBMB nomenclature.In some further embodiments, the natural gum polysaccharide is an xanthan gum.
[06] the present invention also provides cleaning method, comprising: the object that will have natural gum polysaccharide spot contacts with the cleaning compositions that comprises transglucosidase; Under the condition that is enough to finish the degraded of natural gum polysaccharide, keep contacting of object and cleaning compositions, and then cleaning objects.In some embodiments, object is to be besmirched by the food that comprises a kind of natural gum polysaccharide at least.In some further embodiments, the natural gum polysaccharide comprises xanthan gum.Further in the embodiment, object is selected from fabric and hard surface at some.In some other embodiment, transglucosidase has the aminoacid sequence with about at least 90% identity of aminoacid sequence of Aspergillus transglucosidase.In some embodiments, cleaning compositions also comprises at least a tensio-active agent.In some further embodiments, cleaning compositions further comprises at least a enzyme that is used for other spot composition degradeds, is selected from proteolytic enzyme, amylase, cellulase, lipase, cutinases, mannase, polygalacturonase, pectate lyase and oxydo-reductase.And in some other embodiment, cleaning compositions has about 5.0 to about 11.5 pH value.In some further embodiments, cleaning compositions comprises that concentration range is the transglucosidase of about 0.01ppm to about 100ppm.
[07] in some embodiments, the invention provides the composition that comprises transglucosidase and at least a natural gum polysaccharide; Wherein said natural gum polysaccharide is as the substrate of described transglucosidase.In some embodiments, the natural gum polysaccharide comprises xanthan gum and/or guar gum.Yet the present invention is not limited to any certain tree glue polysaccharide.In some preferred embodiments, the natural gum polysaccharide exists as the spot of object (for example fabric), and in other embodiment, it is free in the solution.In some embodiments, transglucosidase comprises at least about 70% and being same as, and is same as at least about 80%, be same as at least about 85%, be same as at least about 90%, be same as at least about 95%, or at least about 98% aminoacid sequence that is same as the aminoacid sequence of Aspergillus transglucosidase.In some embodiments, composition further comprises at least a tensio-active agent and/or other sanitising agents.
[08] the present invention also provides and comprises the method for transglucosidase and at least a natural gum polysaccharide combination with the described natural gum polysaccharide of degrading.In some embodiments, described method comprises: the cleaning compositions that will comprise transglucosidase contacts the object that natural gum polysaccharide spot is arranged; Object and cleaning compositions are remained under the natural gum polysaccharide condition that is enough to effectively degrade together, thus cleaning objects.
[09] in some embodiments, object (for example fabric) is to be made dirty by food, and it comprises at least a natural gum polysaccharide (food that for example, comprises at least a xanthan gum and/or guar gum).In some embodiments, be other spot compositions of degrading, cleaning compositions further comprises a kind of tensio-active agent, other sanitising agents at least, and/or at least a other enzyme (for example, proteolytic enzyme, amylase, cellulase, lipase, cutinase, oxydo-reductase etc.).In some embodiments, the about pH5.0 of pH scope of cleaning compositions is to about pH11.5, and the concentration range that transglucosidase exists in cleaning compositions is that about 0.01ppm is to about 100ppm.
[010] in some embodiments, here the method that is provided is more effectively removed the spot that contains the natural gum polysaccharide than the equivalent means of not using transglucosidase.
[011] in some further embodiments, the invention provides isolating enzyme, it contains the Aspergillus transglucosidase that is produced by Trichodermareesei (Trichoderma reesei) host cell.The present invention also provides the cleaning compositions that the enzyme that provides is provided (for example, cloth-washing detergent), and the method for using enzyme cleaning objects (for example, fabric).
Description of drawings
[012] can understand some aspect of following detailed description in conjunction with the accompanying drawings best.It is emphasized that according to common way each feature of accompanying drawing is not drawn to scale.On the contrary, for clarity sake the size of each feature is at random to enlarge or to reduce.
[013] Fig. 1 provides the collection of illustrative plates of pTrex3 (AGL51M) carrier.
[014] Fig. 2 provides the nucleotide sequence (SEQ ID NO:1) of pTrex3 (AGL51M) expression cassette.
[015] Fig. 3 is the transglucosidase measured with the reducing sugar method figure to the enzymic activity of 0.2% xanthan gum.
[016] Fig. 4 is for being presented in the 50mM Hepes damping fluid (pH7.4) and in AATCC HDL washing composition (pH7.4), the figure of the clean-up performance of little cloth specimen (figure below) that little cloth specimen (microswatches) (last figure) that Trip-TG makes dirty to guar gum and salad dressing are made dirty.
[017] Fig. 5 provides the figure of dose response experiments, and being presented among the AATCC HDL Trip-TG is effective to little cloth specimen that salad dressing is made dirty when 1-5ppm concentration.
[018] Fig. 6 provides a figure, is presented at the cleaning action of the middle Trip-TG of high-performance solid washing composition (HDD) to the little cloth specimen of salad dressing.
[019] Fig. 7 provides a figure, is presented in stirring formula detersive power determinator (Tergotometer) detection, uses 0.15% AATCC HDL, and Trip-TG is to the cleaning action of marmalade spot.
The specific descriptions of invention
[020] the invention provides composition, it comprises transglucosidase and natural gum polysaccharide, wherein the sky Right gummy polysaccharide is the substrate of transglucosidase. The present invention also provides and uses the transglucosidase degraded natural The method of natural gum polysaccharide. In some preferred embodiments, said composition and method find that it is clearly Purposes in the clean application.
[021] before more detailed description exemplary, should be appreciated that the present invention is not limited to Therefore particular described herein can change certainly. Also should be appreciated that art used herein Language only is for describing particular, being not meant to limit the present invention.
[022] for number range used herein, should be understood that except as otherwise noted, when providing one During number range, (its accuracy is to this lower limit unit for the median between its upper and lower bound 1/10th) also by clearly open. Between any setting value each is more among a small circle or at setting range Interior median and any other setting in this setting range or median all are encompassed in invention In. These upper and lower bounds more among a small circle can be comprised in independently in this scope or be excluded at this Outside the scope, and wherein one of the upper and lower bound of each its scope be included in this more among a small circle in, on The limit and lower limit all be not included in this more among a small circle in upper and lower bound all be included in this more among a small circle in Each scope also be encompassed in the present invention, be subjected to any limit of specifically getting rid of in the setting range System. All setting ranges comprise one or two boundaries of boundary, get rid of one or two included boundary Scope also drop in the present invention.
[023] although those similar or be equal to here described any method and material can Use in practice of the present invention or the test, describe now typical method and material. Mention herein All patents and publication be together with being incorporated herein by reference that publication is quoted, with open and Describing method and/or material comprise sequence. It is any that this paper quotes
Figure A20088000933700081
The sequence of database Acceptance integral body is incorporated herein by reference, and comprises wherein nucleic acid and protein sequence and these sequences Note, as this patent the earliest the applying date desired.
[024] only because before their applying date that is disclosed in the application and this paper discussion is provided Publication. Admit that the present invention can be owing to formerly invention prior to these publication but should not be construed as Thing. In addition, the open date that provides may be different from the open date of reality, may need independent confirmation.
[025] except having in addition the regulation at this, under all technology used herein and science vocabulary and the invention The field those of ordinary skill usually understand have an identical meanings. Although find similar or be equal to Described herein those many methods and material can be used for practice of the present invention or the test in, still Some preferred method and materials have been described herein. Number range comprises the numeral of limited range, with And each numeral therebetween.
Definition
[026] used odd number " a ", " an " and " the " wraps herein and in the additional claim Draw together plural number and refer to, unless offer some clarification in addition in the context. Therefore, for example, mention " host cell " The plural form that comprises this host cell.
[027] except as otherwise noted, respectively, nucleic acid be with 5 ' write from left to right to 3 ' direction; Ammonia The base acid sequence with amino to the carboxyl direction from left to right direction write. Title provided herein is not to this The restriction of invention various aspects or embodiment, these embodiments or aspect can be passed through with reference to conduct Whole application documents and obtaining. Therefore, and then following defined term passes through reference as whole The application documents of body and being defined more fully.
[028] term " reorganization " is meant and non-natural is present in polynucleotide or polypeptide in the host cell.In some embodiments, recombinant molecule comprises two or more naturally occurring sequences, and they are being not that naturally occurring mode links together.Reconstitution cell comprises the polynucleotide or the polypeptide of reorganization.
[029] term " heterology " is meant the element that can not be mutually related usually.For example, if host cell produces heterologous protein, then this protein can't help normally that this host cell produces.Equally, effectively being connected to the promotor of one section allogeneic coding sequence, is the promotor that effectively is connected to the encoding sequence that can effectively not be connected to usually in the wild-type host cell.Term " homology " when referring to polynucleotide or protein, is meant naturally occurring polynucleotide or protein in host cell.
[030] term " protein " and " polypeptide " are used interchangeably in this paper context.
[031] " signal sequence " is to be present in proteinic N-terminal amino acids sequence, and it helps the protein maturation form from emiocytosis.The definition of signal sequence is functional definition.The mature form of extracellular protein lacks signal sequence, and this sequence is cleaved in secretion process to be fallen.
[032] " encoding sequence " is the dna segment of coded polypeptide.
[033] term " nucleic acid " comprises DNA, RNA, strand or double-stranded and its chemically modified form.Term " nucleic acid " and " polynucleotide " are used interchangeably in this paper context.
[034] " carrier " refers to be intended to Nucleotide is introduced the polynucleotide of one or more host cells.Suitable carriers is at different host cell self-replicatings, and it comprises: cloning vector, expression vector, shuttle vectors, plasmid, phage particle, box etc.
[035] " expression vector " used herein means DNA construct, and it comprises the protein coding region that effectively is connected to suitable control sequence, and this control sequence can influence protein expression in proper host cell.In some embodiments, this type of control sequence comprises that promotor, control that influence is transcribed transcribes the optional operon sequence that produces mRNA, the sequence that coding mRNA goes up suitable ribosome bind site, and the sequence with translation termination is transcribed in enhanser and other controls.
[036] " promotor " is the regulating and controlling sequence that starts the downstream transcribed nucleic acid.
[037] term " effectively connect " is meant and makes the arrangement of the functional relevant element of element.For example, if promotor control encoding sequence transcribing it effectively is connected to this sequence.
[038] term " selective marker " be meant can be in the host expressed protein, make those contain nucleic acid or the host of carrier is easy to select through introducing.But the example of selective marker includes but not limited to, microbiotic (for example, Totomycin, bleomycin or paraxin) and/or host cell is given the gene of metabolic advantage.The nutritional advantages of host cell for example.
[039] term " is derived " and is comprised that term " is derived from ", and " acquisition ", " can obtain from " and " separating oneself ", and refer to sequence and/or protein source.
[040] " non-virulent " biology is to people and/or the non-pathogenic biology of other animals.
[041] term " recovery ", " separation " and " segregation " are as used herein, finger protein matter, cell, nucleic acid or amino acid with at least a with it natural relevant component separate.
[042] used herein, relate to the term " conversion " that is used for cell, " stable conversion " and " transgenosis " meaning and be phalangeal cell have non-protogenous (for example, heterology) nucleotide sequence, it is incorporated in the cellular genome or as the free plasmid and keeps in many generations.
[043] used herein, term " expression " refers to produce based on the gene nucleic acid sequence process of polypeptide.This process comprises simultaneously to be transcribed and translates.
[044] term " introducing " that uses in the context that nucleotide sequence is inserted in the cell refers to " transfection ", " conversion " or " transduction ", and comprise following implication: nucleic acid is incorporated in eucaryon or the prokaryotic cell prokaryocyte, and wherein this nucleotide sequence can be incorporated into (for example karyomit(e), plasmid, plastid or Mitochondrial DNA) in the genome of this cell, is converted into spontaneous replicon or transient expression (for example mRNA of transfection).
[045] term " hybridization " refers to the process that nucleic acid chains is converged by base complementrity pairing known in the art and its complementary strand.Under highly strict hybridization and wash conditions,, think that nucleic acid and reference nucleic acid sequence are " alternative hybridization " in moderate if two sequence-specifics are hybridized each other.Moderate and height stringent hybridization condition are well known by persons skilled in the art.An example of height stringent condition comprises, at about 42 ℃, 50% methane amide, 5 * SSC, 5 * Denhardt ' s solution, among the 0.5%SDS and 100 μ g/ml modified support DNA hybridize, then in room temperature through 2 * SSC and 0.5%SDS washed twice, and at 42 ℃ with 0.1 * SSC and 0.5%SDS washed twice again.
[046] used herein, " cleaning compositions " and " cleaning formulation " refers to composition, and it can be used for removing undesirable compound (for example, " spot ") from the article that will clean, as fabric, tableware, contact lens, other solid substrates, hair (shampoo), skin (soap and emulsifiable paste), tooth (mouth wash shua, toothpaste) etc.This term comprises that (as long as any material/compound of) particular type cleaning compositions for example, liquid, gel, particle, or spray composition is the main body enzyme compatibility in this composition and the composition through selecting to be used to have needed product form.Considering needs clean Surface, article and/or fabric, and the composition desired form that is used for clean conditions when using, and the concrete selection of cleaning compositions material is easy to make.
[047] this term be intended to include, but are not limited to detergent composition (as liquid, gel, and/or solid laundry detergents, high-count fabric washing composition; The hard surface cleaning preparation, as glass, timber, pottery, the top of metal cabinet and window; Carpet cleaner; The baking box sanitising agent; The fabric freshener; Fabric softener; And weaving and laundry pre-washing agent (pre-spotters), and dish washing detergent).
[048] except as otherwise noted, term used herein " cleaning compositions " comprises, particle, tablet or omnipotent powder type or efficient cleaner, particularly sanitising agent; Liquid, gel or omnipotent lotion form washing composition, the type of particularly so-called efficient liquid (HDL); Liquid high-count fabric washing composition; Manual dish washing detergent or flexible dish washing detergent comprise high expansion type; Dishware detergent comprises the various tablets that family and mechanism use, particle, liquid and rinse aid type; Liquid ceanser and sterilizing agent comprise antimicrobial form hand washing type, cleaning rod, mouth wash shua, artificial tooth detergent, automobile or carpet washing composition, bathroom detergent, shampoo and hair conditioner; Shower gels, foam bath; Metal detergent, and as bleaching washing assistant and " spot rod ", pre-treatment, or the cleaning additive of laundry additive type.
[049] used herein, term " detergent composition " and " detergent formulations " are meant and are used for using with cleaning made dirty object and composition prepared at cleaning medium.In specific embodiments, this term is used in reference to laundering of textile fabrics and/or clothes (as " cloth-washing detergent ").In an alternative embodiment, this term refers to other washing composition, is used for the cleaning disc ware as those, tableware etc. (for example, " dish washing detergent ").The object of the invention is not to limit any specific detergent formulations or composition.In fact, its objective is that except the main body enzyme, cleanser compositions also can comprise tensio-active agent, transferring enzyme, lytic enzyme, oxydo-reductase, washing assistant, SYNTHETIC OPTICAL WHITNER, bleach-activating agent, bluing agent and fluorescence dye, caking inhibitor, sequestering agent, zymoexciter, antioxidant and/or solubilizing agent etc.
[050] used herein, " enhanced performance " is defined as spot (particularly those comprises the relevant spot of natural gum polysaccharide in the cleaning compositions, as milk chocolate, salad sauce, guar-bean etc.) enhancing cleaning (for example, remove and/or decolouring), it is by determining by common evaluation behind the standard wash cycle.
[051] used herein, term " hard-surface cleaning compositions " refers to clean the cleanser compositions as crusts such as floor, wall, ceramic tile, bathroom and galley equipment.This based composition can provide in any form, includes but not limited to solid, liquid, emulsion etc.
[052] used herein, " dish washing compositions " refers to clean the composition of all forms of tableware, includes but not limited to gel, particle and liquid form.
[053] used herein, " the clean fabric composition " is meant the cleanser compositions of all forms of clean textile, includes but not limited to gel, particle, liquid and bar shaped.
[054] used herein, " textiles " refers to woven fabric, and converts to or as the staple fibre and the long filament of yarn, woven, knitting and supatex fabric.The yarn of being made by natural and synthetic (as artificial) fiber contained in this term.
[055] used herein, " textile materials " is the general name of fiber, intermediate yarn, yarn, fabric and the product (for example, clothes and other article) made from fabric.
[056] used herein, " fabric " comprises any textile materials.Therefore, its objective is that this term comprises clothes, and fabric, yarn, fiber, nonwoven material, natural materials, synthetic materials and any other textile materials.
[057] " significant quantity of transglucosidase " used herein refers to reach the amount of the essential transglucosidase of required enzymic activity in concrete application the (as cleaning compositions etc.).This significant quantity determined by one of ordinary skill by this area easily, and is based on many factors, as the enzyme of the specific mutation of use, cleaning applications, the concrete composition of cleaning compositions, and liquid or (for example do, granular, strip) whether composition is essential, or the like.
[058] term " transglucosidase " refers to free or is incorporated into 1, the enzyme of 4-α-D-dextran, and it shifts 1, and the α on 4-α-D-dextran-D-glucosyl is to the one-level hydroxyl of glucose.Transglucosidase as herein described has the enzyme nomenclature according to IUBMB, is described as the activity of EC2.4.1.24.The systematic name of transglucosidase as herein described is 1,4-α-D-dextran: 1, and 4-α-D-dextran (D-glucose) 6-α-D-Transglucosylase.This kind of enzyme can be called alpha-glucosidase in some is open.
[059] term " object of making dirty " refers to second composition object (for example, fabric or dish ware) of (for example, " besmirching ") of making dirty.What contained by term " object of making dirty " is dirty fabric, as soiled clothes, and sheet and by containing the fabric that the natural gum polysaccharide food is polluted.In some embodiments, this pollution manifests the visible color, and in other embodiments, it can not manifest the visible color.
[060] term " natural gum polysaccharide " refers to the non-starch polysaccharide of natural origin, and it can cause soltion viscosity to increase considerably under low concentration.This polysaccharide is generally to be used for foodstuffs industry, and numerous food product (as food flavouring, cream, milk preparation, ice-creams, wood this, milk shake, salad sauce etc.) in, use as thickening material, jelling agent, emulsifying agent and stablizer.Guar gum (foodstuff additive E412), the edible thickening material that from leguminous plants guar-bean shrub, extracts, and xanthan gum (foodstuff additive E415), a kind of by glucose or sucrose (for example, xanthomonas campestris (Xanthomonascampestris)) polysaccharide of fermentation generation is the example of natural gum polysaccharide.Other natural gum polysaccharide include, but are not limited to agar (E406), Lalgine (E400), beta-glucan, carrageenan (E407), tuno gum, Dammar gum, gellan (E418), glucomannan (E425), Sudan Gum-arabic (E414), India(n) gum, tragacanth gum (E413), kuteera gum (E416), locust tree beanpod glue (E410), frankincense gum, sodium alginate (E401), spruce gum and tara gum (E417).
[061] term " non-starch food polysaccharide degrading enzyme " refers to enzyme, the polysaccharide of its non-starchy food of degrading.Typical enzyme includes, but not limited to hemicellulase, mannase, polygalacturonase, zytase, beta-galactosidase enzymes and alpha-galactosidase.
[062] term " working pH value " is meant the pH value in the washing composition use.For example, the working pH value of cloth-washing detergent is the pH value of washing composition when being used for laundering of textile fabrics in washing machine and/or hand washing process.Equally, the working pH value of dish washing detergent is to use in dishwasher and/or the pH value of this washing composition when manual dishwashing.In some embodiments, concentrate or the washing composition of solid form, before the pH of cloth-washing detergent is in its working pH, diluted or dissolving.
[063] term " working concentration " refers to the concentration of enzyme in its use in the cloth-washing detergent.For example, the working concentration of enzyme is the concentration of this enzyme when cloth-washing detergent is used in washing machine and/or hand-washes cleaning fabric in the cloth-washing detergent.Equally, the working concentration of enzyme is to be used in dishwasher and/or the manual dishwashing process concentration of enzyme in this dish washing detergent in dish washing detergent.In some embodiments, the enzyme in washing composition is in before its working concentration, dilutes or dissolve the washing composition of concentrated or solid form.
Detailed Description Of The Invention
[064] the invention provides composition, comprise transglucosidase and natural gum polysaccharide, wherein, the natural gum polysaccharide is the substrate of transglucosidase.The present invention also provides the method for using transglucosidase degraded natural gum polysaccharide.In some preferred embodiments, find that said composition and method can be used for cleaning applications.In fact, the invention provides the multiple composition that contains transglucosidase, and use these method for compositions.
The composition that contains transglucosidase
[065] as mentioned above, the invention provides the composition that contains transglucosidase.In some embodiments, said composition comprises at least a transglucosidase and natural gum polysaccharide, and wherein the natural gum polysaccharide is the substrate of transglucosidase.
[066] as mentioned above, transglucosidase has the activity that is defined as EC2.4.1.24 according to the IUBMB enzyme nomenclature usually, and it is active to be to shift the glucosyl of some dextran to the one-level hydroxyl of glucose.In some embodiments, this enzyme also can have following activity, and it is by cutting sugared side chain or cracking interior keys to destroy the polysaccharide skeleton natural gum polysaccharide (as xanthan gum and contain galactomannan polysaccharide, as guar gum or Lima bean gum) of degrading.
[067] find any suitable transglucosidase can be used for the present invention (for example see people such as Pazur, Carbohydr.Res.1986 149:137-47[1986]; With people such as Nakamura, J.Biotechnol., 53:75-84[1997]).In some embodiments, can be used for transglucosidase of the present invention and be the merchant sells and obtainablely (for example, includes but not limited to from Megazyme Wicklow, Ireland; Or Danisco US Inc., Genencor Division, Palo Alto, CA).In some embodiments, this kind of enzyme is aspergillus niger (Aspergillus niger) transglucosidase that the Trichodermareesei cell produces.In some other embodiment, transglucosidase is that (for example, include but not limited to have the fungi transglucosidase of following aminoacid sequence, this sequence is stored in NCBI's to wild-type fungi transglucosidase
Figure A20088000933700141
In the database, sequence accept number be: D45356 (GID:2645159; Aspergillus niger), BAD06006.1 (GID:4031328; Aspergillus awamori (Aspergillus awamori)), BAA08125.1 (GID:1054565; Aspergillus oryzae (Aspergillus oryzae)), XP_001210809.1 (GID:115492363; Terreus (Aspergillus terreus)), XP_001271891.1 (GID:121707620; Rod aspergillus (Aspergillus clavatus)), XP_001266999.1 (GID:119500484; Neosartorya fischeri), XP_751811.1 (GID:70993928; Aspergillus fumigatus (Aspergillus fumigatus)), XP_659621.1 (GID:67523121; Aspergillus nidulans (Aspergillus nidulans)), XP_001216899.1 (GID:115433524; Terreus (Aspergillus terreus)) and XP_001258585.1 (GID:119473371; Neosartoryafischeri)), or its variant, it has aminoacid sequence with wild-type fungi transglucosidase at least about 70% identity, at least about 80% identity, at least about 85% identity, at least about 90% identity, at least about 95% identity, or at least about the aminoacid sequence of 98% identity.
[068] in some preferred embodiments, this enzyme usually with about 0.01ppm (1,000,000/, w/v) be present in the composition (0.01ppm about 0.05ppm extremely according to appointment to the concentration of about 100ppm, about 0.05ppm is to about 0.1ppm, about 0.1ppm is to about 0.5ppm, and about 0.5ppm is to about 1ppm, and about 1ppm is to about 5ppm, about 5ppm is to about 10ppm, and about 10ppm is to about 100ppm).
[069] in some preferred embodiments, said composition is a cleaning compositions.In some particularly preferred embodiments, said composition further comprises at least a tensio-active agent, clean-out system and/or other fabric care agents, and more detailed description is arranged below.
[070] in some embodiments, the natural gum polysaccharide is in aqueous solution, and in other embodiments, it is attached to (for example, being present in body surface as spot, as fabric or crust) on the object.In some embodiments, the natural gum polysaccharide is the exsiccant form.Because the natural gum polysaccharide generally is used for various food, so in some embodiments, object is to be made dirty by the food that contains the natural gum polysaccharide.The typical food that contains the natural gum polysaccharide include, but are not limited to food flavouring, cream, milk preparation, ice-creams, wood this, milk shake, salad sauce, nectar, tinned fruit, jelly, soybean sauce, oyster sauce, packing meat, cheese, and baked goods.In some embodiments, be present in natural gum polysaccharide concentration in the food and be about 0.1% to about 1.5%, about 0.1% to about 0.5%, about 0.5% to about 1.0%, about 1.0%, or about 1.5%.
[071] in some preferred embodiments, transglucosidase is to adopt produced in conventional processes.For example, in some embodiments, it is (for example by gram-negative micro-organism, as intestinal bacteria (E.coli)) be secreted into pericentral siphon, or (for example by gram-positive microorganism and actinomycetes (Actinomycetes) such as bacillus (Bacillus)) are secreted into extracellular space, or produce by eucaryon host (for example Trichoderma (Trichoderma), Aspergillus, yeast belong (Saccharomyces) and Pichia (Pichia)).
[072] in some embodiments, transglucosidase produces by expressed fusion protein in the Trichodermareesei host cell, and this fusion rotein contains the signal sequence that effectively is connected on the transglucosidase.In some of these embodiments, transglucosidase is secreted in the substratum, and can gather in the crops from substratum.Any signal sequence can be used in the suitable fusion rotein, so that promote protein from the Trichoderma secretory host cell.In some embodiments, signal sequence is endogenic, and in other embodiment, is non-endogenous for the Trichoderma host cell, and in some embodiments, this protein signal sequence is known to be by a large amount of excretory signal sequences from Trichoderma species host cell.The sort signal sequence comprises, but be not limited to: the signal sequence of cellobiohydrolase I, cellobiohydrolase II, endoglucanase i, EG II, EG III, α-Dian Fenmei, aspartyl protease, glucoamylase, mannase, Glycosylase and barley endopeptidase B (is seen, for example, Saarelainen, Appl.Environ.Microbiol., 63:4938-4940[1997]).In some embodiments, and in embodiment of the present disclosure part institute further described, transglucosidase was used its own signal sequence and obtained secreting (being AGL1, AGL2 or AGL3 signal sequence).
[073] therefore, in some embodiments, use following nucleic acid to produce transglucosidase, this nucleic acid comprises the signal sequence encoding nucleic acid that effectively is connected on the transglucosidase coding nucleic acid, wherein this translated nucleic acid produces fusion rotein, this fusion rotein comprises the transglucosidase part, and it has a N-terminal signal sequence from Trichoderma secretory host cell transglucosidase part.
[074] in some embodiments, except that signal sequence, fusion rotein further comprises " carrier proteins ", and it is a proteinic part, and this protein is endogenic for Trichodermareesei species host cell, and by its a large amount of secretions.Suitable carriers protein comprises, but be not limited to Trichodermareesei mannase I (Man5A or MANI), Trichodermareesei cellobiohydrolase II (Cel6A or CBHI1) (for example sees, people such as Paloheimo, Appl.Environ.Microbiol., 69:7073-7082[2003]) or Trichodermareesei cellobiohydrolase I (CBHI) those.In some embodiments, carrier proteins is the Trichodermareesei CBH1 albumen of truncation type, and it comprises CBH1 nucleus and portion C BH1 connecting zone.Therefore, in some embodiments, use the nucleic acid of encoding fusion protein, this fusion rotein comprises from the N-terminal to the C-terminal, signal sequence, carrier proteins and the theme enzyme that effectively is connected.
[075] in some embodiments, carry out codon optimized to the encoding sequence of expressing the transglucosidase of using in the transglucosidase host cell.Because codon use table is listed in the utilization ratio of each codon in many host cells (comprising Trichodermareesei), this is knownly (for example to see in this area, people such as Nakamura, Nucl.Acids Res., 28:292[2000]) or derive easily, the nucleic acid of coding transglucosidase aminoacid sequence to be expressed can be designed so at an easy rate.
[076] in some embodiments, except encoding sequence, nucleic acid has also comprised in host cell expresses other required elements for transglucosidase.For example, in some embodiments, nucleic acid comprises and is used for promotor and the transcription terminator that encoding sequence is transcribed.The promotor that typically is used for Trichodermareesei includes, but are not limited to Trichodermareesei cbh1, cbh2, egl1, egl2, eg5, xln1 and xln2 promotor, or its heterozygosis or truncation type variant.For example, in some embodiments, promotor is a Trichodermareesei cbh1 promotor.The terminator that is fit to comprises Trichodermareesei cbh1, cbh2, egl1, egl2, eg5, xln1 and xln2 terminator and many other, for example comprise terminator from aspergillus niger or Aspergillus awamori glucoamylase gene, as well-known those skilled in the art, and Aspergillus nidulans anthranilic acid synthase gene, aspergillus oryzae TAKA amylase gene, or the trpC of Aspergillus nidulans (people such as Punt, Gene 56:117-124[1987]).In some embodiments, promotor and/or terminator are natural for Trichoderma species host cell, and in other embodiment, all right and wrong are endogenic for Trichoderma species host cell for they.
[077] in some embodiments, wherein the Trichodermareesei host cell is used to express transglucosidase, and this cell carries out genetic modification expresses to reduce the endogenic secretory protein of cell.In some embodiments, cell comprises the gene of one or more natural genes, particularly encoded secreted protein, its deleted or inactivation.For example, in some embodiments, one or more protease-encoding genes (for example, aspartyl encoding gene; See people such as Berka, Gene 86:153-162[1990]; With US Pat.No.6,509,171) or the deleted or inactivation of cellulase encoding gene.In some embodiments, Trichoderma species host cell is to contain deactivation disappearance cbh1, and the Trichodermareesei host cell of cbh2 and egl1 and egl2 gene is described as WO 05/001036.In some embodiments, the nucleic acid of foregoing description is present in the nuclear gene group of Trichoderma species host cell, and in other embodiment, is present in the plasmid that duplicates in the Trichoderma host cell.
[078] anyly (for example finds can be used among the present invention for the appropriate method of in the Trichoderma host cell, introducing nucleic acid, electroporation, the nuclear microinjection, transduction, transfection, the transfection of liposome-mediated and the mediation of DEAE-dextrin is hatched with calcium phosphate DNA precipitation, bombards at a high speed and protoplastis merges with the particulate of DNA bag quilt).In fact, conventional transformation technology be well known in the art (for example see people such as Campbell, Curr.Genet., 16:53-56[1989]; WO 05/001036; US Pat.No.6,022,725; US Pat.No.6,103,490; US Pat No.6,268,328; With the laid-open U.S. Patents application serial no: 20060041113,20060040353,20060040353 and 20050208623, it openly is incorporated herein by reference).In some embodiments, the preparation that is used to the Trichoderma that transforms comprises the preparation protoplastis from radicula byssoidea.(for example see, people such as Campbell, the same).In some embodiments, mycelium is sprouted from plant spore and is obtained.
[079] in some embodiments, in case be secreted in the nutrient solution, this transglucosidase is promptly with any suitable, easily method reclaim (for example, centrifugal, affine by precipitation, filter or any other method well known in the art.For example, find available, affinity chromatography (people such as Tilbeurgh, FEBSLett., 16:215[1984]); Ion exchange chromatography (people such as Goyal, Biores.Technol., 36:37[1991]; People such as Fliess, Eur.J.Appl.Microbiol., Biotechnol.17:314[1983]; People such as Bhikhabhai, J.Appl.Biochem., 6:336[1984]; With people such as Ellouz, Chromatogr., 396:307[1987]), comprise the ion-exchange of using the capable of high resolution material (people such as Medve, J.Chromatogr.A 808:153[1998]; Hydrophobic interaction chromatography (Tomaz and Queiroz, J.Chromatogr.A 865:123[1999]; Two-phase is distributed (people such as Brumbauer, Bioseparation 7:287[1999]); Ethanol sedimentation; RPLC; At silica gel or the chromatography on Zeo-karb (as DEAE), chromatofocusing, SDS-PAGE electrophoresis; Ammonium sulfate precipitation; Or gel-filtration (for example, using sephadex G-75).In some embodiments, use is without the transglucosidase of purifying from other compositions of substratum.In these embodiments some, substratum uses through concentrating simply then, need not to be further purified protein from the composition of growth medium, and in other embodiment, and it uses without any further modification or processing.
[080] the present invention also provides the method for degraded natural gum polysaccharide.In some embodiments, this method is included in and under the condition that makes the natural gum glycocalix degrade transglucosidase is combined (as mixing) with the natural gum polysaccharide.It is known in the art being suitable for the active condition of transglucosidase (as being suitable for the active temperature range of transglucosidase, pH value scope and other reacted constituents).
Cleaning method
[081] except the above-described composition that contains transglucosidase, the present invention also provides cleaning method.These methods generally comprise: the cleaning compositions that comprises transglucosidase contacts with the object of being made dirty by at least a natural gum polysaccharide, be enough to cause under the condition of natural gum polysaccharide degraded, common this object of maintenance and cleaning compositions, thus clean this object.
[082] in some preferred embodiments, used cleaning compositions is clean fabric composition (for example, cloth-washing detergent) in these methods, surface cleaning composition, tableware cleaning compositions, automatic dishwasher detergent composition etc.The several formulations that can be used for cleaning compositions of the present invention, those that include, but are not limited in WO 0001826 to describe in detail very much, it is incorporated herein this paper as a reference.
[083] in some embodiments, this described cleaning compositions (as cloth-washing detergent) (for example comprises at least a about 1% to about 80%, about 5% to about 50% (calculating by weight)) tensio-active agent (for example, nonionogenic tenside, cats product, anion surfactant, and/or zwitterionics, or its any mixture, as the mixture of negatively charged ion and nonionogenic tenside).The typical surface promoting agent includes, but are not limited to: alkylbenzene sulfonate (ABS) comprises linear alkyl benzene sulfonate and linear alkyl sodium sulfonate, the alkyl phenoxy polyethoxyethanols (for example, Nonylphenoxy ethoxylate or nonyl phenol), diethanolamine, trolamine and Monoethanolamine MEA BASF.The exemplary surfactants that can be used for various washing composition, particularly cloth-washing detergent at U.S. Patent number 3664961,3919678, is described in 4222905 and 4239659, and it is incorporated herein by reference separately.
[084] in some embodiments, this described cleaning compositions is solid (for example, the form of powder or tablet), liquid or gel.In some preferred embodiments, composition comprises that further at least a damping fluid (for example, yellow soda ash or sodium bicarbonate), washing auxiliary detergent, SYNTHETIC OPTICAL WHITNER, bleach-activating agent, enzyme, enzyme stabilizers, suds booster, inhibitor, anti-tarnishing agent, corrosion inhibitor, soil-suspending agent, stain remover, sterilant, pH adjust agent, non-washing assistant source of alkalinity, sequestrant, organic or inorganic filler, solvent, expanding material, white dyes, dyestuff and/or perfume.In some embodiments, cleaning compositions combines with washing composition before using as laundry additive.
[085] in some embodiments, for removing other spots, cleaning compositions further comprises at least a non-starchy food polysaccharide degrading enzyme (for example hemicellulase, mannase, polygalacturonase, zytase or pectate lyase), and alternatively, one or more (for example plant other enzymes, proteolytic enzyme is as subtilisin and/or SSI albumen; Lipase, amylase, cellulase, cutinase, lipase, oxydo-reductase etc.).
[086] multiple other compositions that are used for the washing composition cleaning compositions can be used in the composition provided herein, include, but not limited to other activeconstituentss, carrier, expanding material, processing washing assistant, dyestuff or pigment, liquid preparation solvent etc.Increase in the extra embodiment lathery soap lather synergistic agent such as C at needs 10-C 16Alkolamides is included into composition, accounts for about 1% to about 10% of composition weight usually.
[087] in some embodiments, detergent composition comprises that water and other solvents are as carrier.Low-molecular-weight one-level or secondary alcohol, methyl alcohol for example, ethanol, propyl alcohol and Virahol can be used as such carrier.In some embodiments, monohydroxy-alcohol is preferably as the dissolving tensio-active agent, but polyvalent alcohol, as those contain from about 2 to about 6 carbon atoms and from about 2 to about 6 oh groups (for example, 1, ammediol, ethylene glycol, glycerine and 1, the 2-propylene glycol) also available.In some further embodiments, composition comprises about 5% to about 90%, is generally this class carrier of about 10% to about 50%.
[088] in some preferred embodiments, prepare detergent composition provided herein, make that washing water pH value is between about 5.0 and about 11.5 between usage period of in water clean operation.Therefore, the product of finishing is formulated in this scope usually.Control pH value comprises the use of damping fluid, alkali, acid etc. in the technology of recommending usage level, and this is well-known for those skilled in the art.In some embodiments, this cleaning compositions is to clean dish washing detergent automatically, its working pH scope is at the extremely about pH11.5 of about pH9.0, approximately pH9.0 is to about pH9.5, approximately pH9.5 is to about pH10.0, about pH10.0 is pH10.5 extremely approximately, and about pH10.5 is to about pH11.0, or about pH11.0 is to about pH11.5.In other the embodiment, this cleaning compositions is a liquid laundry detergent at some, and its working pH scope is at the extremely about pH8.5 of about pH7.5, and approximately pH7.5 is to about pH8.0, and about pH8.0 is pH8.5 extremely approximately.In other the embodiment, this cleaning compositions is a solid laundry detergents at some, and its working pH scope is at the extremely about pH10.5 of about pH9.5, and the approximately extremely about pH10.0 of pH9.5, or about pH10.0 is pH10.5 extremely approximately.
[089] various bleaching compounds, as percarbonate, perborate and analogous products also can be used for various composition provided herein, generally account for about 1% to about 15% level by weight.If desired, this composition also contains bleach-activating agent, tetraacetyl ethylene diamine for example, the ninth of the ten Heavenly Stems acyloxy Phenylsulfonic acid and its analogue, this also is well-known in the art.Usage level by weight generally about 1% to about 10% scope.
[090] various stain removers, the few lipid type of negatively charged ion particularly, various sequestrants, particularly amido phosphonate and ethylenediamine disuccinic acid, various clay household cleansers (clay soil removalagents), particularly ethoxylation tetren, various dispersion agents, particularly polyacrylic ester and poly aspartic acid, various whitening agent, particularly negatively charged ion whitening agent, various suds suppressors, particularly siloxanes and secondary alcohol, various fabric softeners, particularly montmorillonitic clay, with its analogue, also can be used in the various compositions, by weight, horizontal extent about 1% to about 35%.Standard preparation and disclosed patent comprise a plurality of detailed description to this conventional material that is used for composition provided by the invention.
[091] in some embodiments, enzyme stabilizers also can be used for this cleaning compositions provided herein.This stablizer comprises, but is not limited to propylene glycol (preferably about 1% to about 10%), sodium formiate (preferably from about 0.1% to about 1%) and calcium formiate (preferably from about 0.1% to about 1%).
[092] in some embodiments, hard-surface cleaning compositions and/or clean fabric composition are formulated as and comprise multiple washing assistant, weight level scope from about 5% to about 50%.Typical washing assistant includes, but are not limited to the 1-10 micron zeolite, and polycarboxylate is as Citrate trianion and oxidation disuccinate, layered silicate, phosphoric acid salt etc.Other conventional washing assistants are listed in standard preparation, and find to can be used among the present invention.
[093] other optional compositions comprise sequestrant, clay greasiness removal/anti redeposition agent, polymeric dispersant, SYNTHETIC OPTICAL WHITNER, whitening agent, suds suppressor, solvent and hairdressing agent.
[094] cleaning method provided by the invention is more effectively removed some spot (for example, from the food stain that contains the natural gum polysaccharide) than the same class methods of not using transglucosidase.In some preferred embodiments, do not comprise the equivalent method of transglucosidase with respect to other, it is more effective that cleaning compositions of the present invention is removed spot.For example, use the standard detection based on reflexometer, this described method is removed than the equivalent means of not using transglucosidase and/or is discolored about at least 20%, at least about 40%, about at least 60%, about at least 80%, or in certain embodiments, more spots of about at least 90%.
[095] in order to further specify the present invention and its advantage, provide following specific embodiment, should understand that these embodiment are provided is for the present invention is described, and the scope that should not be construed as limiting the invention by any way.
Experiment
[096] provides following examples so that how to make and use complete open and description of the present invention for those those of ordinary skills provide, but and do not mean that it is the restriction of the scope of the invention that the contriver is thought, do not mean that following experiment representative is all or the experiment of only finishing yet.For the accuracy of guaranteeing relevant numeral (as quantity, temperature etc.) use aspect is made an effort, but should consider some testing errors and deviation.Unless otherwise indicated, part is by weight a part, and molecular weight is a weight average molecular weight, and temperature is degree centigrade, and pressure is or near normal atmosphere.
Embodiment 1
The expression of aspergillus niger transglucosidase in the Trichodermareesei
[097] nucleic acid of the ripe transglucosidase of coding aspergillus niger by pcr amplification from the aspergillus niger genomic dna and be cloned into the pTrex3 carrier and obtain pTrex3 (AGL51M).PTrex3 (AGL51M) as shown in Figure 1.The transglucosidase albumen of vector encoded effectively is connected to the CBH1 signal sequence thus, is secreted in the growth medium to assist it.There are Trichodermareesei cbh1 promotor and terminator in the both sides of transglucosidase encoding sequence.The nucleotides sequence of pTrex3 (AGL51M) vector expression box is listed in Fig. 2.
[098] the 7.57kb XbaI-XbaI fragment of plasmid pTrex3 (AGL5IM) transforms the Trichodermareesei spore through the agarose gel electrophoresis purifying and by electroporation.The electroporation parameter is as follows: voltage-16kV/cm, electric capacity-25 μ F, resistance-50 Ω.Implement electroporation and use the Trichodermareesei spore suspension that is suspended in the fresh results in the ice-cold 1.2M sorbyl alcohol.Behind the electroporation, spore in substratum on shaking table overnight incubation (300 ℃, 200rpm), substratum contains the 1M sorbyl alcohol, 0.3% glucose, 0.3%Bacto peptone and 0.15% yeast extract.The spore of sprouting is planted in selective medium, and it contains ethanamide as only nitrogen source (ethanamide 0.6g/L; Cesium chloride 1.68g/L; Glucose 20g/L; Potassium primary phosphate 15g/L; Magnesium sulfate heptahydrate 0.6g/L; Calcium dichloride dihydrate 0.6g/L; Ferric sulfate (II) 5mg/L, zinc sulfate 1.4mg/L, cobalt chloride (II) 1mg/L; Manganous sulfate (II) 1.6mg/L; Agar 20g/L, pH4.25).The transformant bacterium colony occurred in about 1 week.Independent transformant is transferred to new ethanamide selectivity flat board, and allows growth 3-4 days.
[099] chorista that shows stable growth on the selective medium is seeded in the invisible spectro 5ml lactose of the 20 * 175mm restricted type substratum (for example seeing 2005/001036, the 60 page of WO).Test tube is fixed in the rotary shaker, with about 45, the 200rpm jolting, 28 ℃ 4-5 days.Electrophoretic analysis to culture supernatant shows to exist to have new protein band about as the expection molecular weight.
[0100] the transglucosidase activity of transformant generation also uses Enzymology method to determine.Detection is carried out in 100mM sodium-acetate buffer (pH4.5), and it contains 4mM p-nitrophenyl-alpha-glucosaccharase and 1mg/mL bovine serum albumin.30 ℃ hatched through 30 minutes after, add equal-volume 1M yellow soda ash termination reaction and also write down OD 405Generally, the transformant of every milliliter of nutrient solution is expressed the transglucosidase activity (being expressed as micromole's amount of the p-nitrophenol of per minute release) of 1-2U.In unconverted contrast, activity is lower than detectability.
Embodiment 2 transglucosidases degraded xanthan gum
[0101] as known in the art, enzyme uses PAHBAH (P-hydroxybenzoic acid hydrazides) reagent to measure (see people such as Lever, Anal.Biochem., 47:273[1972]) through the reducing sugar detection method to the xanthan gum hydrolytic activity.Xanthan gum (CAS 11138-66-2) is available from Sigma Chemicals, and St.LouisMO also is dissolved in the 50mM sodium-acetate buffer (pH6.0), and concentration is 0.2%.For some experiment AATCC standard efficient liquid washing composition (the AATCC HDL 2003 of no whitening agent, TestFabrics, Inc.West Pittston, PA) every liter of adding 1.5ml (0.15%).Contain 12% linear alkyl sulfonate in this AATCC HDL liquid washing agent, 8% ethanol oxyethyl group, 8% propylene glycol, 1.2% citric acid, 4% lipid acid and 4% sodium hydroxide, and by water balance.
[0102] (COSTAR 3526 in 24 hole microwell plates, Corning Incorporated, Corning, NY) damping fluid that carries out following detection: 1ml joins hole 1, the enzyme-added hole 2 that joins of the damping fluid of 1ml, the damping fluid of 1ml and substrate join the enzyme-added and substrate of the damping fluid of hole 3 and 1ml and join hole 4.Be the statistics purpose, every hole can be provided with 2 to 4 times.Enzyme to be detected dilutes with 1 to 10 to 1 to 1000 in reaction buffer usually.After adding all reagent, place plastic cover on the microwell plate and lid and plate junction with Parafilm closely several layers of parcels (Pechiney PlasticPacking, Menasha, WI), to avoid evaporating.Sptting plate is at 37 ℃, and 100rpm rotational vibration was hatched 1 to 16 hour.
[0103] Eppendorf Mastercycler Gradient (Eppendorf Scientific is used in the reducing sugar determination of activity, Westbury, NY) the disposable PCR of thermal cycler and 0.2ml (polymerase chain reaction) bar pipe and lid are available from VWR International, West Chester, PA.Reducing sugar reagent is prepared as follows: 10ml 2% sodium hydroxide is dissolved in distilled water, adds 0.15g four water sodium-potassium tartrates (Rochelle Salt, Sigma Chemical Co.) and 0.15g P-hydroxybenzoic acid hydrazides (H-9882, Sigma Chemical Co.).This solution (being called " PAHBAH reagent ") vortex is with the dissolving all the components, and is placed on and places the dark place to using on ice.Fresh preparation every day of this reagent.Before the analysis, 0.160ml PAHBAH reagent joins in each PCR bar pipe, adds enzyme sample and the contrast of 5 to 20 μ l subsequently.All pipes cover lid completely, place thermal cycler, and hatch 15 minutes at 99 ℃, 4 ℃ of coolings subsequently at least 15 minutes.After the cooling, remove the bar pipe cap, get each sample of 0.15m and place the flat microwell plate in 96 holes (COSTAR 9017, Corning Inc.Corning, NY), with Spectra Max 250Plate Reader (Molecular Devices, Sunnyvale, CA) at the 405nm reading, distilled water is blank.
[0104] each enzyme sample carries out following analysis: the optical density(OD) of control sample (OD) is deducted from the OD of buffer sample, and this value is added in the substrate buffer solution contrast.Enzyme adds the OD of substrate reactions liquid and the summation of substrate and sample contrast compares.
[0105] Fig. 3 shows, the transglucosidase that Trichoderma (Trip-TG) and aspergillus (Mega-TG) produce adds to demonstrate for xanthan gum in the AATCC efficient liquid washing composition at 50mM acetate buffer solution (pH6.0) significant reducing sugar activity.
Embodiment 3 transglucosidases are removed the spot on the dyeing patch
[0106] the cotton cloth specimen of making dirty by the salad sauce that contains pigment (STC CFT CS-6) and guar-bean pigment (STC CFT CS-43), available from Test Fabrics, Inc.West Pittston, PA, USA.Detect for ease of microwell plate, " the weaving Punch Press Model B of die-cutting machine cuts into 15mm circle (disk) to cloth specimen with being equipped with 5/8.Single cloth specimen disk is placed every hole of 24 hole microwell plates (Costar 3526).One (1) milliliter scavenging solution joins in each hole, and its every liter contains 1.5ml AATCCHDL washing composition, 50mM Hepes damping fluid and be diluted in 6 in the 50mM pH7.4Hepes damping fluid to the 60ppm enzyme.Microwell plate covers with plastic cover and aluminium foil parcel and the gentle rotation of 100rpm, hatches 4-16 hour for 37 ℃.Plate takes out from vibrator and the sucking-off detergent solution.Each hole of microwell plate is inferior with 1.5ml Dulbecco ' s PBS pH7.3 washing three (3), and 1.5ml distilled water wash three (3) is inferior.Each disk shifts out from its hole, and between paper handkerchief dried overnight, avoid directly being exposed to light.The visual inspection disk is analyzed with Minolta Reflectometer CR-200 then, and it is calibrated with the color standard white ceramic tile.Calculating has the average L value of standard deviation value.
[0107] figure that provides of Fig. 4 shows that Trip-TG (being diluted in the 50mM Hepes damping fluid with 1/50) adds at 50mM pH7.4Hepes damping fluid and 50mM pH7.4Hepes damping fluid and can remove salad sauce spot and guar-bean pigment spot among the 0.15%AATCC HDL.
[0108] Fig. 5 shows available from the little cloth specimen result of experiment of Trip-TG dose response.In the 0.15%AATCC efficient liquid, 1ppm Trip-TG has removed the salad sauce spot.
[0109] Fig. 6 shows, in the experiment of little cloth specimen cleaning, Trip-TG removes the salad sauce spot in not containing the 0.1%NorthAmerican AATCC-1993HDD standard of whitening agent.This North American AATCC-1993HDD contains 18% linear alkyl sulfonate, 25% zeolite A, 18% soda, 0.5% water glass, 22% sodium sulfate, 10% moisture and 6.3% multipolymer or other additives.
[0110] this experiment is presented at that transglucosidase all has the greasiness removal activity in efficient liquid (HDL) and the high-performance solid (HDD).
The stirring formula detersive power determinator of embodiment 4 transglucosidase cleaning actions is analyzed
[0111] stir the research of formula detersive power determinator use 6 jars of maintaining 30 ℃ stir formula detersive power determinator 7243S types (US Testing, Co.Inc.Hoboken, NJ).Stirring velocity is set at 100rpm.Cotton sample (each stir in formula detersive power determinator jar 5) is available from Warwick EquestLimited, Consett, County Durham, England, to there be the cotton sample of food stain circle to join 1 liter of 0.15%AATCC HDL washing composition, contain 6gpg hardness (from containing the 15000gpg hardness stock solution dilution of 1.735M calcium chloride and 0.67M magnesium chloride) and 25mM pH7.4HEPES damping fluid.Through 30 minute wash(ing)cycle, cloth specimen dried 7 minutes and removes unnecessary water with 1.5L cold running water flushing three times, and dried overnight under the room temperature.After reflectometer is analyzed each stain, calculate spot with standard method and discharge per-cent (%SRI).Fig. 7 shows, transglucosidase with do not contain the enzyme contrast or contain irrelevant protein, bovine serum albumin (BSA-50, component V does not contain immunoglobulin (Ig) and proteolytic enzyme, available from Rockland Immunochemicals, Gilbertsville, PA.) contrast is compared, and can significantly clean the jam stain.
[0112] above embodiment shows, the transglucosidase xanthan gum of degrading effectively, and remove some spot from the cotton sample.
[0113] all patents mentioned in the specification sheets and the level that makes a declaration of one of ordinary skill in the art of the present invention.All patents with open with quote each respectively and separately and disclose identical degree and quote this paper as a reference.
[0114] those skilled in the art are easy to understand, and the present invention is very suitable for realizing purpose of the present invention, and result who obtains mentioning and advantage, and those advantages of institute's inherent wherein.The composition described herein and the method for description are exemplary, represent embodiment preferred, rather than limitation of the scope of the invention.Can change, replace and revise and not deviate from scope and spirit of the present invention the present invention's content disclosed herein and it will be apparent to those skilled in the art that.
[0115] invention described herein can be implemented under the situation that does not have the concrete disclosed element of those this paper, restriction.Term of Shi Yonging and statement herein only is used for the description rather than the restriction of term, and the use of these terms and statement is not intended to get rid of, and any this place is showed or equivalent or its part of the feature of description, but it should be understood that within the scope of the invention, can carry out various modifications.Therefore, should be realized that, though by feature is specifically open in the preferred specific embodiments and optionally, the modification of theory disclosed herein and variant can be used by those skilled in the art, and these are revised and variant falls within the scope of the present invention in present invention.
[0116] the present invention extensively and has synoptically been described here.Drop on open each interior narrower range of generality and the next classification and also form a part of the present invention.This comprises generality description of the present invention that has restriction or the opposition restriction of getting rid of any theme from sort out, and whether the material of no matter getting rid of is quoted especially at this paper.

Claims (19)

1. composition comprises:
A) transglucosidase; With
B) natural gum polysaccharide;
Wherein said natural gum polysaccharide is the substrate of described transglucosidase.
2. the composition of claim 1, wherein said natural gum polysaccharide is an xanthan gum.
3. the composition of claim 1, wherein said transglucosidase has the activity that is defined as EC2.4.1.24 according to the IUBMB nomenclature.
4. the composition of claim 1, wherein said transglucosidase have and the amino acid of the Aspergillus transglucosidase aminoacid sequence at least about 90% identity.
5. the composition of claim 1, wherein said composition further comprises at least a tensio-active agent.
6. the composition of claim 1, wherein said natural gum polysaccharide exists as the spot on the object.
7. the composition of claim 6, wherein said object is a fabric.
8. a method comprises transglucosidase and natural gum polysaccharide combined with the described natural gum polysaccharide of degrading.
9. the method for claim 8, wherein said transglucosidase has the activity that is defined as EC2.4.1.24 according to the IUBMB nomenclature.
10. the method for claim 8, wherein said natural gum polysaccharide is an xanthan gum.
11. a cleaning method comprises:
A) object that the natural gum polysaccharide is made dirty contacts with the cleaning compositions that comprises transglucosidase; With
B) under the described natural gum polysaccharide condition that is enough to effectively to degrade, keep described object and cleaning compositions, thereby clean described object.
12. the cleaning method of claim 11, wherein said object are to be made dirty by the food that contains the natural gum polysaccharide.
13. the cleaning method of claim 11, wherein said natural gum polysaccharide comprises xanthan gum.
14. the cleaning method of claim 11, wherein said object is selected from fabric and crust.
15. the cleaning method of claim 11, wherein said transglucosidase have and the amino acid of the Aspergillus transglucosidase aminoacid sequence at least about 90% identity.
16. the cleaning method of claim 11, wherein said cleaning compositions further comprises at least a tensio-active agent.
17. the cleaning method of claim 11, wherein said cleaning compositions also comprises at least a enzyme, and it is selected from proteolytic enzyme, amylase, cellulase, lipase, cutinases, mannase, polygalacturonase, pectate lyase and oxydo-reductase are with other spot compositions of degrading.
18. the cleaning method of claim 11, the pH of wherein said cleaning compositions are that about pH5.0 is to about pH11.5.
19. the cleaning method of claim 11, wherein said cleaning compositions comprise that concentration range is at the described enzyme of about 0.01ppm to about 100ppm.
CN200880009337A 2007-03-22 2008-03-21 The cleaning compositions that contains transglucosidase Pending CN101641433A (en)

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CN113834757B (en) * 2021-07-30 2024-02-23 广东美味鲜调味食品有限公司 Rapid detection method for high-temperature-resistant amylase trace residue of oyster sauce raw material

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