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1. WO2017177153 - METHODS FOR SELECTING ENZYMES HAVING PROTEASE ACTIVITY

Publication Number WO/2017/177153
Publication Date 12.10.2017
International Application No. PCT/US2017/026626
International Filing Date 07.04.2017
IPC
C12N 15/10 2006.01
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
10Processes for the isolation, preparation or purification of DNA or RNA
CPC
C12N 15/1058
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
10Processes for the isolation, preparation or purification of DNA or RNA
1034Isolating an individual clone by screening libraries
1058Directional evolution of libraries, e.g. evolution of libraries is achieved by mutagenesis and screening or selection of mixed population of organisms
C12N 15/1075
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
10Processes for the isolation, preparation or purification of DNA or RNA
1034Isolating an individual clone by screening libraries
1075by coupling phenotype to genotype, not provided for in other groups of this subclass
C12Q 1/37
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
QMEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS
1Measuring or testing processes involving enzymes, nucleic acids or microorganisms
34involving hydrolase
37involving peptidase or proteinase
C40B 20/02
CCHEMISTRY; METALLURGY
40COMBINATORIAL TECHNOLOGY
BCOMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES, IN SILICO LIBRARIES
20Methods specially adapted for identifying library members
02Identifying library members by their fixed physical location on a support or substrate
C40B 40/08
CCHEMISTRY; METALLURGY
40COMBINATORIAL TECHNOLOGY
BCOMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES, IN SILICO LIBRARIES
40Libraries per se, e.g. arrays, mixtures
04Libraries containing only organic compounds
06Libraries containing nucleotides or polynucleotides, or derivatives thereof
08Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
C40B 50/06
CCHEMISTRY; METALLURGY
40COMBINATORIAL TECHNOLOGY
BCOMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES, IN SILICO LIBRARIES
50Methods of creating libraries, e.g. combinatorial synthesis
06Biochemical methods, e.g. using enzymes or whole viable microorganisms
Applicants
  • NOVOZYMES A/S [DK]/[DK]
  • BLAZEJ, Robert [US]/[US] (BW)
Inventors
  • BLAZEJ, Robert
  • TORIELLO, Nicholas
  • EMRICH, Charles
Agents
  • LAMBIRIS, Elias
  • FECHTER, Eric
Priority Data
62/319,74707.04.2016US
Publication Language English (EN)
Filing Language English (EN)
Designated States
Title
(EN) METHODS FOR SELECTING ENZYMES HAVING PROTEASE ACTIVITY
(FR) PROCÉDÉS DE SÉLECTION D'ENZYMES À ACTIVITÉ PROTÉASIQUE
Abstract
(EN)
Provided herein are systems and components thereof for improving protease activity. The systems make use of an emulsion for in vitro compartmentalization of a library of synthetic compounds, each compound having a gene linked to a protease substrate and selectable marker. Expressed enzymes with greater protease activity will preferentially hydrolyze the protease substrate, whereas enzymes with less protease activity will leave the substrate intact. Removal of the non-hydrolyzed compounds provides an enriched gene library encoding for more active protease variants. Also described are synthetic compounds and emulsions which can be used in the methods.
(FR)
L'invention concerne des systèmes et des composants de ces derniers pour améliorer l'activité protéasique. Les systèmes utilisent une émulsion pour la compartimentation in vitro d'une bibliothèque de composés synthétiques, chaque composé ayant un gène lié à un substrat de protéase et un marqueur sélectionnable. Les enzymes exprimées présentant une plus forte activité protéasique vont préférentiellement hydrolyser le substrat des protéases, tandis que des enzymes présentant une activité protéasique moindre laisseront le substrat intact. L'élimination des composés non hydrolysés permet d'obtenir une bibliothèque enrichie par des gènes codant pour des variants de protéases plus actifs. L'invention concerne également des composés synthétiques et des émulsions qui peuvent être utilisés dans lesdites méthodes.
Also published as
Latest bibliographic data on file with the International Bureau