title: Directed evolution of squalene synthase for dehydrosqualene biosynthesis.
abstract: Squalene synthase (SQS) catalyzes the first step of sterol/hopanoid biosynthesis in various organisms. It has been long recognized that SQSs share a common ancestor with carotenoid synthases, but it is not known how these enzymes selectively produce their own product. In this study, SQSs from yeast, human, and bacteria were independently subjected to directed evolution for the production of the C30 carotenoid backbone, dehydrosqualene. This was accomplished via high-throughput screening with Pantoea ananatis phytoene desaturase, which can selectively convert dehydrosqualene into yellow carotenoid pigments. Genetic analysis of the resultant mutants revealed various mutations that could effectively convert SQS into a "dehydrosqualene synthase." All of these mutations are clustered around the residues that have been proposed to be important for NADPH binding.
authors.affiliation: Department of Applied Chemistry and Biotechnology, Chiba University, 1-33 Yayoi-cho, Inage, Chiba 263-8522, Japan.
authors.forename: Maiko
authors.initials: M
authors.lastname: Furubayashi
authors.affiliation: Department of Applied Chemistry and Biotechnology, Chiba University, 1-33 Yayoi-cho, Inage, Chiba 263-8522, Japan.
authors.forename: Ling
authors.initials: L
authors.lastname: Li
authors.affiliation: Department of Applied Chemistry and Biotechnology, Chiba University, 1-33 Yayoi-cho, Inage, Chiba 263-8522, Japan.
authors.forename: Akinori
authors.initials: A
authors.lastname: Katabami
authors.affiliation: Department of Applied Chemistry and Biotechnology, Chiba University, 1-33 Yayoi-cho, Inage, Chiba 263-8522, Japan.
authors.forename: Kyoichi
authors.initials: K
authors.lastname: Saito
authors.affiliation: Department of Applied Chemistry and Biotechnology, Chiba University, 1-33 Yayoi-cho, Inage, Chiba 263-8522, Japan; Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (JST), 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Umeno
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics*; D055162:Biocatalysis; D019020:Directed Molecular Evolution; D013186:Farnesyl-Diphosphate Farnesyltransferase / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D008958:Models, Molecular; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics*; D013185:Squalene / Q000031:analogs & derivatives* / Q000138:chemical synthesis
chemical_list: D001426:Bacterial Proteins; D029701:Saccharomyces cerevisiae Proteins; C000864:dehydrosqualene; D013185:Squalene; D013186:Farnesyl-Diphosphate Farnesyltransferase
keywords: Carotenoid synthase; Directed evolution; Phytoene desaturase; Squalene synthase