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WO2019110492A1 - Cellule de levure recombinante - Google Patents

Cellule de levure recombinante Download PDF

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Publication number
WO2019110492A1
WO2019110492A1 PCT/EP2018/083321 EP2018083321W WO2019110492A1 WO 2019110492 A1 WO2019110492 A1 WO 2019110492A1 EP 2018083321 W EP2018083321 W EP 2018083321W WO 2019110492 A1 WO2019110492 A1 WO 2019110492A1
Authority
WO
WIPO (PCT)
Prior art keywords
promoter
prk
recombinant yeast
yeast cell
expression
Prior art date
Legal status (The legal status 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 status listed.)
Ceased
Application number
PCT/EP2018/083321
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English (en)
Inventor
Ioannis PAPAPETRIDIS
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
DSM IP Assets BV
Original Assignee
DSM IP Assets BV
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 DSM IP Assets BV filed Critical DSM IP Assets BV
Publication of WO2019110492A1 publication Critical patent/WO2019110492A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/10—Transferases (2.)
    • C12N9/12—Transferases (2.) transferring phosphorus containing groups, e.g. kinases (2.7)
    • C12N9/1205—Phosphotransferases with an alcohol group as acceptor (2.7.1), e.g. protein kinases
    • 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
    • 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
    • C12N9/88—Lyases (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
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
    • C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
    • C12P7/06—Ethanol, i.e. non-beverage
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y207/00—Transferases transferring phosphorus-containing groups (2.7)
    • C12Y207/01—Phosphotransferases with an alcohol group as acceptor (2.7.1)
    • C12Y207/01019—Phosphoribulokinase (2.7.1.19)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y401/00—Carbon-carbon lyases (4.1)
    • C12Y401/01—Carboxy-lyases (4.1.1)
    • C12Y401/01039—Ribulose-bisphosphate carboxylase (4.1.1.39)
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
    • Y02E50/00—Technologies for the production of fuel of non-fossil origin
    • Y02E50/10—Biofuels, e.g. bio-diesel

Definitions

  • the invention relates to a recombinant yeast cell having the ability to produce a desired fermentation product, to the functional expression of heterologous peptides in a yeast cell, and to a method for producing a fermentation product wherein said yeast cell is used.
  • mutated as used herein regarding proteins or polypeptides means that at least one amino acid in the wild-type or naturally occurring protein or polypeptide sequence has been replaced with a different amino acid, inserted or deleted from the sequence via mutagenesis of nucleic acids encoding these amino acids.
  • Mutagenesis is a well-known method in the art, and includes, for example, site-directed mutagenesis by means of PCR or via oligonucleotide-mediated mutagenesis as described in Sambrook et al., Molecular Cloning-A Laboratory Manual, 2nd ed., Vol. 1-3 (1989).
  • nucleic acid variations are "silent variations" and represent one species of conservatively modified variation.
  • PRK is brought under control of a weak constitutive promotor, the cells were able to grow under aerobic conditions, but when such cell were use in ethanol fermentation, there was still significant amount of glycerol production.
  • neither strong, medium strong, or weak constitutive promotors are feasible for PRK with the purpose to reduce glycerol formation.
  • a functionally expressed phosphoribulokinase (PRK, (EC 2.7.1.19)) according to the invention is capable of catalyzing the chemical reaction:
  • the fermentable carbohydrate is obtained from starch, lignocellulose, and/or pectin.
  • the organic compound made with the process of the invention is selected from the group consisting of ethanol, n-butanol, 2-butanol, isobutanol, lactic acid, 3-hydroxy- propionic acid, acrylic acid, acetic acid, succinic acid, fumaric acid, malic acid, itaconic acid, maleic acid, citric acid, adipic acid, an amino acid, such as lysine, methionine, tryptophan, threonine, and aspartic acid, 1 ,3-propane-diol, ethylene, glycerol, a b-lactam antibiotic and a cephalosporin, vitamins, pharmaceuticals, animal feed supplements, specialty chemicals, chemical feedstocks, plastics, solvents, fuels, including biofuels and biogas or organic polymers, and an industrial enzyme, such as a protease, a cellulase, an amylase, a glucanase, a lactase,
  • the DAN1 gene of S cerevisiae is regulated in parallel with the hypoxic gene, but by a different mechanism, 1997, Gene Vol 192, pag 199-205.

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Genetics & Genomics (AREA)
  • Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Mycology (AREA)
  • Medicinal Chemistry (AREA)
  • Plant Pathology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

La présente invention concerne une cellule de levure recombinante exprimant fonctionnellement une ou plusieurs séquences d'acides nucléiques hétérologues codant pour la ribulose-1,5-phosphate carboxylase/oxygénase (EC4.1.1.39 ; RuBisCO), et éventuellement une ou plusieurs chaperonnes moléculaires pour la RuBisCO, et une ou plusieurs phosphoribulokinases (EC2.7.1.19 ; PRK), où la PRK se trouve sous le contrôle d'un activateur de transcription (TA), de sorte que la PRK est située en aval d'un premier promoteur, lequel premier promoteur est induit par ladite TA, et l'expression de ladite TA étant induite par un second promoteur. Le second promoteur peut permettre l'expression supérieure sous des conditions anaérobies par rapport aux conditions aérobies, préférablement il présente un rapport d'expression du TA (anaérobie/aérobie) de 2 ou plus. Le second promoteur peut permettre l'expression lorsque le rapport cellulaire (NADH/NAD+)anaérobie/(NADH/NAD+)aérobie est de 2 ou plus.
PCT/EP2018/083321 2017-12-08 2018-12-03 Cellule de levure recombinante Ceased WO2019110492A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
EP17206139.2 2017-12-08
EP17206139 2017-12-08

Publications (1)

Publication Number Publication Date
WO2019110492A1 true WO2019110492A1 (fr) 2019-06-13

Family

ID=60654779

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/EP2018/083321 Ceased WO2019110492A1 (fr) 2017-12-08 2018-12-03 Cellule de levure recombinante

Country Status (1)

Country Link
WO (1) WO2019110492A1 (fr)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2023285280A1 (fr) * 2021-07-12 2023-01-19 Dsm Ip Assets B.V. Cellule de levure recombinante
WO2023285281A1 (fr) * 2021-07-12 2023-01-19 Dsm Ip Assets B.V. Cellule de levure recombinée
CN115820708A (zh) * 2022-09-06 2023-03-21 厦门大学 感知真菌信息素的重组酿酒酵母菌的构建方法及生物传感器

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008041840A1 (fr) 2006-10-02 2008-04-10 Dsm Ip Assets B.V. Génie métabolique de cellules de levure induisant la fermentation de l'arabinose
WO2009013159A2 (fr) 2007-07-23 2009-01-29 Dsm Ip Assets B.V. Enzymes productrices d'acétyl-coa dans la levure
WO2009112472A2 (fr) 2008-03-13 2009-09-17 Dsm Ip Assets B.V. Sélection d’organismes capables de faire fermenter des substrats mélangés
WO2011010923A1 (fr) 2009-07-24 2011-01-27 Technische Universiteit Delft Production fermentative d'éthanol exempt de glycérol
WO2014129898A2 (fr) 2013-02-22 2014-08-28 Technische Universiteit Delft Microorganisme recombiné destiné à être utilisé dans un procédé avec un rendement en produit accru

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008041840A1 (fr) 2006-10-02 2008-04-10 Dsm Ip Assets B.V. Génie métabolique de cellules de levure induisant la fermentation de l'arabinose
WO2009013159A2 (fr) 2007-07-23 2009-01-29 Dsm Ip Assets B.V. Enzymes productrices d'acétyl-coa dans la levure
WO2009112472A2 (fr) 2008-03-13 2009-09-17 Dsm Ip Assets B.V. Sélection d’organismes capables de faire fermenter des substrats mélangés
WO2011010923A1 (fr) 2009-07-24 2011-01-27 Technische Universiteit Delft Production fermentative d'éthanol exempt de glycérol
WO2014129898A2 (fr) 2013-02-22 2014-08-28 Technische Universiteit Delft Microorganisme recombiné destiné à être utilisé dans un procédé avec un rendement en produit accru

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2023285280A1 (fr) * 2021-07-12 2023-01-19 Dsm Ip Assets B.V. Cellule de levure recombinante
WO2023285281A1 (fr) * 2021-07-12 2023-01-19 Dsm Ip Assets B.V. Cellule de levure recombinée
CN115820708A (zh) * 2022-09-06 2023-03-21 厦门大学 感知真菌信息素的重组酿酒酵母菌的构建方法及生物传感器

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