WO1995020039A3 - Customized proteases with altered transacylation activity - Google Patents

Customized proteases with altered transacylation activity Download PDF

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Publication number
WO1995020039A3
WO1995020039A3 PCT/US1995/006682 US9506682W WO9520039A3 WO 1995020039 A3 WO1995020039 A3 WO 1995020039A3 US 9506682 W US9506682 W US 9506682W WO 9520039 A3 WO9520039 A3 WO 9520039A3
Authority
WO
WIPO (PCT)
Prior art keywords
proteases
transacylation
customized
methods
amino acids
Prior art date
Application number
PCT/US1995/006682
Other languages
French (fr)
Other versions
WO1995020039A2 (en
WO1995020039B1 (en
Inventor
Klaus Breddam
Morten C Kielland-Brandt
Uffe Hasbo Mortensen
Kjeld Ove Olesen
Henning Ralf Stennicke
Fred W Wagner
Original Assignee
Calsberg A S
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Calsberg A S filed Critical Calsberg A S
Priority to AU26513/95A priority Critical patent/AU679861B2/en
Priority to JP7517672A priority patent/JPH09504438A/en
Priority to NZ283995A priority patent/NZ283995A/en
Priority to EP95921432A priority patent/EP0723585A1/en
Publication of WO1995020039A2 publication Critical patent/WO1995020039A2/en
Publication of WO1995020039A3 publication Critical patent/WO1995020039A3/en
Publication of WO1995020039B1 publication Critical patent/WO1995020039B1/en

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/102—Mutagenizing nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/60—Growth-hormone releasing factors (GH-RF) (Somatoliberin)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P21/00—Preparation of peptides or proteins
    • C12P21/02—Preparation of peptides or proteins having a known sequence of two or more amino acids, e.g. glutathione

Abstract

The invention provides customized proteases (i.e., mutant enzymes), e.g. carboxypeptidase Y, methods of making customized proteases, as well as methods of using customized proteases. The customized proteases of the invention are derived from the known proteases. Altered transacylation reactions include the capability to perform transacylation reactions not substantially catalyzed by the known protease or the capability to perform transacylation reactions with improved yields, or both. The methods of the invention provide for customized proteases through site specific or random mutagenesis of the active site amino acids of the known proteases. The invention also provides for methods of using the customized proteases to prepare a preselected transacylation products. The preselected transacylation products produced can be modified by substitution at the N- or C-terminal with nucleophiles such as L-amino acids, D-amino acids, amino acid amides, and radioactive amino acids.
PCT/US1995/006682 1993-10-28 1994-10-28 Customized proteases with altered transacylation activity WO1995020039A2 (en)

Priority Applications (4)

Application Number Priority Date Filing Date Title
AU26513/95A AU679861B2 (en) 1993-10-28 1994-10-28 Customized proteases with altered transacylation activity
JP7517672A JPH09504438A (en) 1993-10-28 1994-10-28 Special protease with modified acyl transfer activity
NZ283995A NZ283995A (en) 1993-10-28 1994-10-28 Customized (mutant) carboxypeptidase Y having modified S1 subsite
EP95921432A EP0723585A1 (en) 1993-10-28 1994-10-28 Customized proteases

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US14470493A 1993-10-28 1993-10-28
US08/144,704 1994-10-27
US08/329,892 1994-10-27
US08/329,892 US6187579B1 (en) 1993-10-28 1994-10-27 Customized proteases

Publications (3)

Publication Number Publication Date
WO1995020039A2 WO1995020039A2 (en) 1995-07-27
WO1995020039A3 true WO1995020039A3 (en) 1996-03-21
WO1995020039B1 WO1995020039B1 (en) 1996-05-30

Family

ID=26842276

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1995/006682 WO1995020039A2 (en) 1993-10-28 1994-10-28 Customized proteases with altered transacylation activity

Country Status (7)

Country Link
US (2) US6187579B1 (en)
EP (1) EP0723585A1 (en)
JP (1) JPH09504438A (en)
AU (1) AU679861B2 (en)
CA (1) CA2174525A1 (en)
NZ (1) NZ283995A (en)
WO (1) WO1995020039A2 (en)

Families Citing this family (42)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB9323429D0 (en) * 1993-11-12 1994-01-05 Wellcome Found Therapy
NZ315713A (en) * 1995-08-16 1999-08-30 Zeneca Ltd A conjugate comprising an antibody as targeting moiety linked to a carboxypeptidase b (cpb)
DE19535082A1 (en) 1995-09-21 1997-03-27 Henkel Ecolab Gmbh & Co Ohg Paste-like detergent and cleaning agent
US6537776B1 (en) 1999-06-14 2003-03-25 Diversa Corporation Synthetic ligation reassembly in directed evolution
US20030219752A1 (en) * 1995-12-07 2003-11-27 Diversa Corporation Novel antigen binding molecules for therapeutic, diagnostic, prophylactic, enzymatic, industrial, and agricultural applications, and methods for generating and screening thereof
US6562594B1 (en) 1999-09-29 2003-05-13 Diversa Corporation Saturation mutagenesis in directed evolution
US6171820B1 (en) 1995-12-07 2001-01-09 Diversa Corporation Saturation mutagenesis in directed evolution
US6238884B1 (en) 1995-12-07 2001-05-29 Diversa Corporation End selection in directed evolution
US6939689B2 (en) 1995-12-07 2005-09-06 Diversa Corporation Exonuclease-mediated nucleic acid reassembly in directed evolution
US20020164580A1 (en) * 1995-12-07 2002-11-07 Diversa Corporation Combinatorial screening of mixed populations of organisms
US20040023327A1 (en) * 1995-12-07 2004-02-05 Short Jay M. End selection in directed evolution
US6361974B1 (en) 1995-12-07 2002-03-26 Diversa Corporation Exonuclease-mediated nucleic acid reassembly in directed evolution
US6713279B1 (en) 1995-12-07 2004-03-30 Diversa Corporation Non-stochastic generation of genetic vaccines and enzymes
US5830696A (en) 1996-12-05 1998-11-03 Diversa Corporation Directed evolution of thermophilic enzymes
US6764835B2 (en) * 1995-12-07 2004-07-20 Diversa Corporation Saturation mutageneis in directed evolution
US7018793B1 (en) 1995-12-07 2006-03-28 Diversa Corporation Combinatorial screening of mixed populations of organisms
US6740506B2 (en) 1995-12-07 2004-05-25 Diversa Corporation End selection in directed evolution
US6352842B1 (en) 1995-12-07 2002-03-05 Diversa Corporation Exonucease-mediated gene assembly in directed evolution
US6358709B1 (en) 1995-12-07 2002-03-19 Diversa Corporation End selection in directed evolution
DE19636035A1 (en) 1996-09-05 1998-03-12 Henkel Ecolab Gmbh & Co Ohg Paste-like detergent and cleaning agent
CA2269659A1 (en) * 1996-11-04 1998-05-14 Merck Frosst Canada & Co. Hydrolase binding assay
DE19703364A1 (en) 1997-01-30 1998-08-06 Henkel Ecolab Gmbh & Co Ohg Paste-like detergent and cleaning agent
US5985627A (en) * 1997-02-28 1999-11-16 Carlsberg Laboratory Modified carboxypeptidase
US6908757B1 (en) 1998-03-26 2005-06-21 The Procter & Gamble Company Serine protease variants having amino acid deletions and substitutions
US6461834B1 (en) * 1998-11-06 2002-10-08 Bionebraska, Inc. Clostripain catalyzed amidation of peptides
DE19857687A1 (en) 1998-12-15 2000-06-21 Henkel Ecolab Gmbh & Co Ohg Pasty detergent
US20090130718A1 (en) * 1999-02-04 2009-05-21 Diversa Corporation Gene site saturation mutagenesis
JP2003505067A (en) 1999-07-22 2003-02-12 ザ、プロクター、エンド、ギャンブル、カンパニー Subtilisin protease variants with amino acid deletions and substitutions in specific epitope regions
CZ2002212A3 (en) 1999-07-22 2002-07-17 The Procter & Gamble Company Variants of subtilisin protease having amino acid substitutions in defined epitope regions
US6946128B1 (en) 1999-07-22 2005-09-20 The Procter & Gamble Company Protease conjugates having sterically protected epitope regions
CZ2002171A3 (en) 1999-07-22 2002-06-12 The Procter & Gamble Company Proteinase conjugate, cleansing agent and preparation for personal hygiene
CN1100789C (en) * 2000-01-13 2003-02-05 中国人民解放军第二军医大学 Preparation process of gene recomibination calcitonin or calcitonin analog
US20030170846A1 (en) * 2000-04-24 2003-09-11 Weisgerber David John Enzyme variants having one or more D-amino acid substitutions
ATE321778T1 (en) * 2000-09-04 2006-04-15 Cellpep Sa MAUROTOXIN, PI1 AND HSTX1 DERIVATIVES
US20040009498A1 (en) * 2002-01-14 2004-01-15 Diversa Corporation Chimeric antigen binding molecules and methods for making and using them
US20030219854A1 (en) * 2002-03-21 2003-11-27 Micrologix Biotech Inc. Methods for producing modified anti-infective peptides
US20040171070A1 (en) * 2002-05-20 2004-09-02 Ramagauri Bhikhabhai Peptide analysis using a solid support
DE602005008071D1 (en) * 2004-08-13 2008-08-21 Hoffmann La Roche C-TERMINAL MODIFICATION OF POLYPEPTIDES
DK1794294T3 (en) * 2004-09-27 2011-10-31 Sanofi Aventis Deutschland Recombinant carboxypeptidase B
EP1979370B1 (en) * 2006-01-17 2010-03-17 N.V. Organon SELECTIVE ENZYMATIC HYDROLYSIS OF C-TERMINAL tert-BUTYL ESTERS OF PEPTIDES
WO2011018421A1 (en) * 2009-08-10 2011-02-17 Morphosys Ag Novel screening strategies for the identification of binders
EP2847316A1 (en) 2012-05-11 2015-03-18 Novozymes A/S A brewing method

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0085516A2 (en) * 1982-01-22 1983-08-10 Carlsberg Biotechnology Ltd. A/S A process for enzymatic replacement of the B-30 amino acid in insulins
WO1992002615A1 (en) * 1990-08-09 1992-02-20 Genentech, Inc. Serine protease variants having peptide ligase activity

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
RO80806A (en) 1979-04-06 1983-02-01 De Forenede Bryggerier As,Dk PROCEDURE FOR ENZYMATIC OBTAINING PEPTIDES
US5185258A (en) 1984-05-29 1993-02-09 Genencor International, Inc. Subtilisin mutants
US4959312A (en) * 1985-05-31 1990-09-25 The University Of Tennessee Research Corporation Full spectrum mutagenesis
JPH07121222B2 (en) 1987-03-24 1995-12-25 寶酒造株式会社 Chemically modified serine protease-containing composition, its production method and use
GB8807271D0 (en) 1988-03-26 1988-04-27 Preece A W Cell fusion apparatus
US5049656A (en) 1988-12-21 1991-09-17 Board Of Regents Of The University Of Nebraska Sequential peptide and oligonucleotide syntheses using immunoaffinity techniques

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0085516A2 (en) * 1982-01-22 1983-08-10 Carlsberg Biotechnology Ltd. A/S A process for enzymatic replacement of the B-30 amino acid in insulins
WO1992002615A1 (en) * 1990-08-09 1992-02-20 Genentech, Inc. Serine protease variants having peptide ligase activity

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
J. BONGERS: "Comparison of enzymatic semisynthesis of peptide amides: human growth hormone releasing factor and analogs", BIOMED. BIOCHIM. ACTA, vol. 50, no. 10/11, pages 157 - 162 *
J. WINTHER ET AL: "Increased hydrophobicity of the S1' binding site in carboxypeptidase Y obtained by site-directed mutagenesis", CARLSBERG RES. COMMUN., vol. 50, pages 273 - 284 *
K OLESEN AND M. KIELLAND-BRANDT: "Altering substrate preference of carboxypeptidase Y by a novel strategy of mutagenesis eliminating wild type background", PROTEIN ENGINEERING, vol. 6, no. 4, pages 409 - 415 *
R. SIEZEN ET AL: "Engineering of the substrate-binding region of the subtilisin-like cell-envelope proteinase of Lactococcus lactis", PROTEIN ENGINEERING, vol. 6, no. 8, pages 927 - 937 *
U. MORTENSEN ET AL: "Site-directed mutagenesis on (serine) carboxypeptidase Y", BIOCHEMISTRY, vol. 33, pages 508 - 517 *

Also Published As

Publication number Publication date
US6187579B1 (en) 2001-02-13
CA2174525A1 (en) 1995-07-27
WO1995020039A2 (en) 1995-07-27
US5945329A (en) 1999-08-31
NZ283995A (en) 2000-04-28
AU679861B2 (en) 1997-07-10
EP0723585A1 (en) 1996-07-31
AU2651395A (en) 1995-08-08
JPH09504438A (en) 1997-05-06

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