# US 10,308,971 B2 · Microorganism for expressing a human membrane protein > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,308,971 B2 - Document ID: US10308971B2 - Status: Granted - Kind: B2 - Application number: 15/891,263 - Filed: 2018-02-07 - Published: 2018-06-21 - Granted: 2019-06-04 - Earliest claimed: 2009-05-21 - Assignees: Novozymes A/S - Applicants: Novozymes A/S - Inventors: Michael Schilling; Christine Lang; Andreas Raab - Web page: https://patentgrep.com/patent/US10308971B2 ## CPC - C12P 33/00: Preparation of steroids - C07K 14/705: Receptors; Cell surface antigens; Cell surface determinants {(tumour specific antigens C07K14/4748)} - C12N 15/81: for yeasts - C12P 21/02: having a known sequence of two or more amino acids, e.g. glutathione - C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)} - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract The invention relates to an isolated, genetically modified, living non-mammal organism, having increased HMG-CoA-reductase activity compared to the wild type, and having reduced C24-methyltransferase and/or delta22-desaturase activity compared to the wild type. The invention is characterized in that the organism has increased dehydrocholesterol-delta70-reductase activity compared to the wild type. The invention further relates to different uses of such an organism, to a test kit comprising such an organism, and to a membrane extract of such an organism. ## Claims ### Claim 1 A method making a membrane protein of a mammal, the method comprising: cultivating an isolated genetically modified living non-mammal yeast organism, having an increased HMG-CoA reductase activity compared to the wild type, and having a reduced C24 methyltransferase and/or delta22 desaturase activity compared to the wild type, wherein the organism has an increased dehydrochole-sterol-delta7 reductase activity compared to the wild type and is additionally transformed with a gene coding for a membrane protein of a mammal; and isolating the membrane protein from the cultivation supernatant and/or from the yeast. ### Claim 2 (depends on 1) The method of claim 1, wherein the organism additionally has an increased dehydrocholesterol-delta24 reductase activity compared to the wild type. ### Claim 3 (depends on 1) The method of claim 1, wherein the organism additionally has an increased squalene epoxidase and/or lanosterol demethylase activity compared to the wild type. ### Claim 4 (depends on 1) The method of claim 1, wherein the organism is selected from the group consisting of Saccharomyces cerevisiae, Saccharomyces delbruckii, Saccharomyces italicus, Saccharomyces ellipsoideus, Saccharomyces fermentati, Saccharomyces kluyveri, Saccharomyces krusei, Saccharomyces lactis, Saccharomyces marxianus, Saccharomyces microellipsoides, Saccharomyces montanus, Saccharomyces norbensis, Saccharomyces oleaceus, Saccharomyces paradoxus, Saccharomyces pastorianus, Saccharomyces pretoriensis, Saccharomyces rosei, Saccharomyces rouxii, Saccharomyces uvarum, Saccharomycodes ludwigii, Kluyveromyces lactis, Kluyveromyces marxianus var. marxianus, Kluyveromyces thermotolerans, Candida utilis, Candida tropicalis, Candida albicans, Candida lipolytica, Candida versatilis, Pichia stipidis, Pichia pastoris, Pichia sorbitophila, and yeasts of the genus Cryptococcus, Debaromyces, Hansenula, Saccharomycecopsis, Saccharomycodes, Schizosaccharomyces, Wickerhamia, Debayomyces, Hanseniaspora, Kloeckera, Zygosaccharomyces, Ogataea, Kuraishia, Komagataella, Metschnikowia, Williopsis, Nakazawaea, Cryptococcus, Torulaspora, Bullera, Rhodotorula, Willopsis and Sporobolomyces. ### Claim 5 (depends on 1) The method of claim 1, wherein the organism is Saccharomyces cerevisiae. ### Claim 6 (depends on 1) The method of claim 1, wherein the organism produces an increased content of desmosterol and/or cholesterol and/or 7-dehydro-cholesterol and/or lathosterol compared to the wild type. ### Claim 7 (depends on 1) The method of claim 1, wherein gene coding for a membrane protein is under the control of a constitutively active promoter. ### Claim 8 (depends on 1) The method of claim 1, wherein cultivating the organism is conducted without addition of cholesterol or desmosterol. ### Claim 9 (depends on 1) The method of claim 1, wherein the organism is cultivated aerobically. ### Claim 10 (depends on 1) The method of claim 1, wherein the membrane protein is a human membrane protein. ### Claim 11 (depends on 1) The method of claim 1, wherein the membrane protein is a plasma membrane protein.