Evolva Sa

University of California

Danmarks Tekniske Universitet

Fluxome Sciences A/s

Evola Sa

Carlsberg A/s

Jochen Förster

Michael Katz

Jens Bredal Nielsen

Hans Peter Smits

Hans Peter Schmidt

Bernhard O Palsson

Imandokht Famili

Hao Luo

Thomas Durhuus

Pengcheng Patrick Fu

Helga David

Malin Sendelius

Bo Stenhuus

Sara Peterson Bjorn

Ross Fennessy

Claes Gjermansen

Jens Bredel Nielsen

Nina Katarina Gunnarsson

Thomas Thomasen Durhuus

Sara Peterson Bjørn

Anna Chailyan

Described herein are recombinant microbial host cells comprising biosynthetic pathways and their use in producing oxidation products and downstream products, e.g., melatonin and related compounds, as well as enzyme variants, nucleic acids, vectors and methods useful for preparing and using such cells. In specific aspects, the present invention relates to monooxygenases, e.g., amino acid hydroxylases, with a modified cofactor-dependency, and to enzyme variants and microbial cells providing for an improved supply of cofactors.

Optimized microbial cells for production of melatonin and other compounds

A recombinant micro-organism producing resveratrol by a pathway in which phenylalanine ammonia lyase (PAL) produces trans-cinnamic acid from phenylalanine, cinnamate 4-hydroxylase (C4H) produces 4-coumaric acid from said trans-cinnamic acid, 4-coumarate-CoA ligase (4CL) produces 4-coumaroyl CoA from said 4-coumaric acid, and resveratrol synthase (VST) produces said resveratrol from said 4-coumaroyl CoA, or in which L-phenylalanine- or tyrosine-ammonia lyase (PAL/TAL) produces 4-coumaric acid, 4-coumarate-CoA ligase (4CL) produces 4-coumaroyl CoA from said 4-coumaric acid, and resveratrol synthase (VST) produces said resveratrol from said 4-coumaroyl CoA. The micro-organism may be a yeast, fungus or bacterium including, or

Metabolically engineered cells for the production of resveratrol or an oligomeric or glycosidically-bound derivative thereof

A recombinant micro-organism such aswhich produces and excretes into culture medium a stilbenoid metabolite product when grown under stilbenoid production conditions, which expresses in above native levels a ABC transporter which transports said stilbenoid out of said micro-organism cells to the culture medium. The genome of theproduces an auxotrophic phenotype which is compensated by a plasmid which also expresses one or more of said enzymes constituting said metabolic pathway producing said stilbenoid, an expression product of the plasmid is genetically modified to include a ubiquitination tag sequence. Expression of an enzyme participating in catabolism of phenylalanine by the Ehrlich pathway is optionally reduced compared to its native expression level.

Production of metabolites

A genetically engineered micro-organism having an operative metabolic pathway producing cinnamoyl-CoA and producing pinosylvin therefrom by the action of a stilbene synthase is used for pinosylvin production. Said cinnamic acid may be formed from L-phenylalanine by a L-phenylalanine ammonia lyase (PAL) which is one accepting phenylalanine as a substrate and producing cinammic acid therefrom, preferably such that if the PAL also accepts tyrosine as a substrate and forms coumaric acid therefrom, the ratio Km(phenylalanine)/Km(tyrosine) for said PAL is less than 1:1 and if said micro-organism produces a cinammate-4 -hydroxylase enzyme (C4H), the ratio K(PAL)/K(C4H) is at least 2:1.

Metabolically engineered cells for the production of pinosylvin

A cis- or trans-stilbenoid of the general formula (1): in which each of R, R, R, Rand Ris hydrogen or hydroxy, or a glycosylated or oligomeric form thereof, is produced by cultivating a micro-organism producing said stilbenoid, in a multi-phase system comprising at least an aqueous first phase containing said micro-organism and a second phase immiscible with said aqueous phase in which (e.g. as which) said stilbenoid accumulates. The second phase may be said stilbenoid or a free or encapsulated solvent compatible with the growth of the micro-organism, for instance an ester. 

Microbial bioreaction process

A genetically engineered micro-organism having an operative metabolic pathway producing cinnamoyl-CoA and producing pinosylvin therefrom by the action of a stilbene synthase is used for pinosylvin production. Said cinnamic acid may be formed from L-phenylalanine by a L-phenylalanine ammonia lyase (PAL) which is one accepting phenylalanine as a substrate and producing cinammic acid therefrom, preferably such that if the PAL also accepts tyrosine as a substrate and forms coumaric acid therefrom, the ratio Km(phenylalanine)/Km(tyrosine) for said PAL is less than 1:1 and if said micro-organism produces a cinammate-4-hydroxylase enzyme (C4H), the ratio K(PAL)/K(C4H) is at least 2:1.

The invention provides an in silico model for determining aphysiological function. The model includes a data structure relating a plurality ofreactants to a plurality ofreactions, a constraint set for the plurality ofreactions, and commands for determining a distribution of flux through the reactions that is predictive of aphysiological function. A model of the invention can further include a gene database containing information characterizing the associated gene or genes. The invention further provides methods for making an in silicomodel and methods for determining aphysiological function using a model of the invention.

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The present invention relates to the construction and engineering of cells, more particularly microorganisms for producing PUFAs with four or more double bonds from non-fatty acid substrates through heterologous expression of an oxygen requiring pathway. The invention especially involves improvement of the PUFA content in the host organism through fermentation optimization, e.g. decreasing the temperature and/or designing an optimal medium, or through improving the flux towards fatty acids by metabolic engineering, e.g. through over-expression of fatty acid synthases, over-expression of other enzymes involved in biosynthesis of the precursors for PUFAs, or codon optimization of the heterologous genes, or expression of heterologous enzymes involved in the biosynthesis of the precursor for PUFAs.

Growing community of inventors

Copenhagen, Denmark

Jochen Förster