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WO2000006718A3 - Expression of functional eukaryotic proteins - Google Patents

Expression of functional eukaryotic proteins Download PDF

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Publication number
WO2000006718A3
WO2000006718A3 PCT/US1999/017127 US9917127W WO0006718A3 WO 2000006718 A3 WO2000006718 A3 WO 2000006718A3 US 9917127 W US9917127 W US 9917127W WO 0006718 A3 WO0006718 A3 WO 0006718A3
Authority
WO
WIPO (PCT)
Prior art keywords
improved
expression
enzymes
relates
eukaryotic proteins
Prior art date
Application number
PCT/US1999/017127
Other languages
French (fr)
Other versions
WO2000006718A2 (en
Inventor
Frances H Arnold
Zhanglin Lin
Original Assignee
California Inst Of Techn
Frances H Arnold
Zhanglin Lin
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 California Inst Of Techn, Frances H Arnold, Zhanglin Lin filed Critical California Inst Of Techn
Priority to IL14050999A priority Critical patent/IL140509A0/en
Priority to EP99935983A priority patent/EP1100891A2/en
Priority to AU51345/99A priority patent/AU5134599A/en
Priority to KR1020017000036A priority patent/KR20010083086A/en
Priority to JP2000562500A priority patent/JP2003503005A/en
Priority to CA002331777A priority patent/CA2331777A1/en
Priority to MXPA01000224A priority patent/MXPA01000224A/en
Publication of WO2000006718A2 publication Critical patent/WO2000006718A2/en
Publication of WO2000006718A3 publication Critical patent/WO2000006718A3/en

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    • 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
    • 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
    • C12N15/1034—Isolating an individual clone by screening libraries
    • C12N15/1058—Directional evolution of libraries, e.g. evolution of libraries is achieved by mutagenesis and screening or selection of mixed population of organisms
    • 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
    • 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
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/67—General methods for enhancing the expression
    • 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
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/80—Vectors or expression systems specially adapted for eukaryotic hosts for fungi
    • C12N15/81—Vectors or expression systems specially adapted for eukaryotic hosts for fungi for yeasts
    • C12N15/815—Vectors or expression systems specially adapted for eukaryotic hosts for fungi for yeasts for yeasts other than Saccharomyces
    • 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
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0065—Oxidoreductases (1.) acting on hydrogen peroxide as acceptor (1.11)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
    • C12Q1/28—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase involving peroxidase
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
    • C07K2319/02—Fusion polypeptide containing a localisation/targetting motif containing a signal sequence
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/795—Porphyrin- or corrin-ring-containing peptides
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/795—Porphyrin- or corrin-ring-containing peptides
    • G01N2333/80—Cytochromes
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/90—Enzymes; Proenzymes
    • G01N2333/902—Oxidoreductases (1.)
    • G01N2333/90245—Oxidoreductases (1.) acting on paired donors with incorporation of molecular oxygen (1.14)

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Plant Pathology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Mycology (AREA)
  • Analytical Chemistry (AREA)
  • Immunology (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Bioinformatics & Computational Biology (AREA)
  • Ecology (AREA)
  • Medicinal Chemistry (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

This invention relates to the improved expression of evolved polynucleotide and polypeptide sequences encoding for eukaryotic enzymes, particularly peroxidase enzymes, in conventional or facile expression systems. Various methods for directed evolution of polynucleotide sequences can be used to obtain the improved sequences. The improved characteristics of the polypeptides or proteins generated in this manner include improved folding, without formation of inclusion bodies, and retained functional activity. In a particular embodiment, the invention relates to improved expression of the horseradish peroxidase gene and horseradish peroxidase enzymes.
PCT/US1999/017127 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins WO2000006718A2 (en)

Priority Applications (7)

Application Number Priority Date Filing Date Title
IL14050999A IL140509A0 (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins
EP99935983A EP1100891A2 (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins
AU51345/99A AU5134599A (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins
KR1020017000036A KR20010083086A (en) 1998-07-28 1999-07-28 Expression of Functional Eukaryotic Proteins
JP2000562500A JP2003503005A (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins
CA002331777A CA2331777A1 (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins
MXPA01000224A MXPA01000224A (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins.

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US9440398P 1998-07-28 1998-07-28
US60/094,403 1998-07-28
US24723299A 1999-02-09 1999-02-09
US09/247,232 1999-02-09

Publications (2)

Publication Number Publication Date
WO2000006718A2 WO2000006718A2 (en) 2000-02-10
WO2000006718A3 true WO2000006718A3 (en) 2000-06-15

Family

ID=26788829

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1999/017127 WO2000006718A2 (en) 1998-07-28 1999-07-28 Expression of functional eukaryotic proteins

Country Status (8)

Country Link
EP (1) EP1100891A2 (en)
JP (1) JP2003503005A (en)
KR (1) KR20010083086A (en)
AU (1) AU5134599A (en)
CA (1) CA2331777A1 (en)
IL (1) IL140509A0 (en)
MX (1) MXPA01000224A (en)
WO (1) WO2000006718A2 (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7166466B2 (en) 1997-12-08 2007-01-23 California Institute Of Technology Method for creating polynucleotide and polypeptide sequences
US7288375B2 (en) 1994-02-17 2007-10-30 Maxygen, Inc. Methods for generating polynucleotides having desired characteristics by iterative selection and recombination

Families Citing this family (20)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6399383B1 (en) 1997-10-28 2002-06-04 Maxygen, Inc. Human papilloma virus vectors
US6596539B1 (en) 1997-10-31 2003-07-22 Maxygen, Inc. Modification of virus tropism and host range by viral genome shuffling
US6902918B1 (en) 1998-05-21 2005-06-07 California Institute Of Technology Oxygenase enzymes and screening method
AU4441699A (en) 1998-06-17 2000-01-05 Maxygen, Inc. Method for producing polynucleotides with desired properties
AU2000275763A1 (en) * 2000-03-27 2001-10-08 California Institute Of Technology Expression of functional eukaryotic proteins
FR2810339B1 (en) * 2000-06-14 2004-12-10 Hoechst Marion Roussel Inc COMBINATORIAL BANKS IMPROVED BY RECOMBINATION IN YEAST AND METHOD OF ANALYSIS
JP2002199890A (en) * 2000-10-23 2002-07-16 Inst Of Physical & Chemical Res Method for modifying biodegradable polyester synthase
US7465567B2 (en) 2001-04-16 2008-12-16 California Institute Of Technology Peroxide-driven cytochrome P450 oxygenase variants
US7226768B2 (en) 2001-07-20 2007-06-05 The California Institute Of Technology Cytochrome P450 oxygenases
EP1639091B1 (en) 2003-06-17 2012-12-05 California University Of Technology Regio- and enantioselective alkane hydroxylation with modified cytochrome p450
EP1660646B1 (en) 2003-08-11 2014-12-31 California Institute Of Technology Thermostable peroxide-driven cytochrome p450 oxygenase variants and methods of use
US11214817B2 (en) 2005-03-28 2022-01-04 California Institute Of Technology Alkane oxidation by modified hydroxylases
US8715988B2 (en) 2005-03-28 2014-05-06 California Institute Of Technology Alkane oxidation by modified hydroxylases
EP2069513A4 (en) 2006-08-04 2012-03-21 California Inst Of Techn METHODS AND SYSTEMS FOR SELECTIVE FLUORATION OF ORGANIC MOLECULES
US8252559B2 (en) 2006-08-04 2012-08-28 The California Institute Of Technology Methods and systems for selective fluorination of organic molecules
US8802401B2 (en) 2007-06-18 2014-08-12 The California Institute Of Technology Methods and compositions for preparation of selectively protected carbohydrates
WO2013016207A2 (en) 2011-07-22 2013-01-31 California Institute Of Technology Stable fungal cel6 enzyme variants
RU2646098C2 (en) * 2013-04-30 2018-03-01 Интас Фармасьютикалс Лтд New method of cloning, expression and purification for preparation of ranibizumab
CA3089002A1 (en) * 2018-01-19 2019-07-25 Obi Pharma, Inc. Crm197 protein expression
CN112852800B (en) * 2020-01-09 2023-02-03 中国科学院天津工业生物技术研究所 A kind of signal peptide mutant and its application

Citations (3)

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Publication number Priority date Publication date Assignee Title
WO1989003424A1 (en) * 1987-10-08 1989-04-20 British Bio-Technology Limited Synthetic gene
WO1997020078A1 (en) * 1995-11-30 1997-06-05 Maxygen, Inc. Methods for generating polynucleotides having desired characteristics by iterative selection and recombination
WO1998006283A1 (en) * 1996-08-14 1998-02-19 Elisabeth Aguilar Apron

Patent Citations (3)

* Cited by examiner, † Cited by third party
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WO1989003424A1 (en) * 1987-10-08 1989-04-20 British Bio-Technology Limited Synthetic gene
WO1997020078A1 (en) * 1995-11-30 1997-06-05 Maxygen, Inc. Methods for generating polynucleotides having desired characteristics by iterative selection and recombination
WO1998006283A1 (en) * 1996-08-14 1998-02-19 Elisabeth Aguilar Apron

Non-Patent Citations (10)

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Title
A. KUCHNER AND F.H. ARNOLD: "Directed evolution of enzyme catalysts", TREND IN BIOTECHNOLOGY, vol. 15, no. 12, December 1997 (1997-12-01), ELSEVIER SCIENCE LTD., AMSTERDAM, NL, pages 523 - 530, XP002133872 *
CRAMERI A ET AL: "IMPROVED GREEN FLUORESCENT PROTEIN BY MOLECULAR EVOLUTION USING DNA SHUFFLING", NATURE BIOTECHNOLOGY,US,NATURE PUBLISHING, vol. 14, 14 March 1996 (1996-03-14), pages 315 - 319, XP000791095, ISSN: 1087-0156 *
F.H. ARNOLD AND J.C. MOORE: "Optimizing industrial enzymes by directed evolution", ADV. BIOCHEM. ENG. BIOTECHNOL., vol. 58, 1997, SPRINGER-VERLAG, BERLIN HEIDELBERG, FRG, pages 1 - 14, XP000891392 *
H. ZHAO AND F.H. ARNOLD: "Combinatorial protein design: strategies for screening protein libraries", CURRENT OPINION IN STRUCTURAL BIOLOGY, vol. 7, 1997, CURRENT BIOLOGY, PHILADELPHIA,US, pages 480 - 485, XP002133873 *
J.C. MOORE ET AL.: "Strategies for the in vitro evolutilon of protein function: enzyme evolution by random recombination of improved sequences", J. MOL. BIOL., vol. 272, no. 3, 26 September 1997 (1997-09-26), ACADEMIC PRESS,US, pages 336 - 347, XP000892797 *
JOO H ET AL: "A high-throughput digital imaging screen for the discovery and directed evolution of oxygenases.", CHEMISTRY AND BIOLOGY, (1999 OCT) 6 (10) 699-706. JOURNAL CODE: CNA., 10 September 1999 (1999-09-10), XP000892823 *
JOO H ET AL: "Laboratory evolution of peroxide-mediated cytochrome P450 hydroxylation [see comments].", NATURE, (1999 JUN 17) 399 (6737) 670-3. JOURNAL CODE: NSC., XP002133874 *
LIN Z ET AL: "Functional expression of horseradish peroxidase in bacteria through directed evolution.", 217TH NATIONAL MEETING OF THE AMERICAN CHEMICAL SOCIETY;ANAHEIM, CALIFORNIA, USA; MARCH 21-25, 1999, vol. 217, no. 1-2, 1999, Abstracts of Papers American Chemical Society 1999, pages BIOT 079, XP000891382, ISSN: 0065-7727 *
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S.A. ORTLEPP ET AL.: "Expression and characterisation of a protein specified by a synthetic horseradish peroxidase gene in Escherichia coli", J. BIOTECHNOLOGY, vol. 11, 1989, ELSEVIER, AMSTERDAM,NL, pages 353 - 364, XP002133871 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7288375B2 (en) 1994-02-17 2007-10-30 Maxygen, Inc. Methods for generating polynucleotides having desired characteristics by iterative selection and recombination
US7166466B2 (en) 1997-12-08 2007-01-23 California Institute Of Technology Method for creating polynucleotide and polypeptide sequences

Also Published As

Publication number Publication date
JP2003503005A (en) 2003-01-28
CA2331777A1 (en) 2000-02-10
AU5134599A (en) 2000-02-21
MXPA01000224A (en) 2002-10-17
IL140509A0 (en) 2002-02-10
WO2000006718A2 (en) 2000-02-10
EP1100891A2 (en) 2001-05-23
KR20010083086A (en) 2001-08-31

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