title: Biochemical characterization of a type III polyketide biosynthetic gene cluster from Streptomyces toxytricini.
abstract: A type III polyketide biosynthetic gene cluster has been discovered in the industrially important strain Streptomyces toxytricini NRRL 15443, including four genes stp450-1, stts, stp450-2, and stmo. The stts gene encodes a putative type III polyketide synthase that is homologous to RppA, a 1,3,6,8-tetrahydroxynaphthalene (THN) synthase from Streptomyces griseus. The deduced protein product of stmo resembles the cupin-containing monooxygenase MomA from Streptomyces antibioticus that oxidizes THN into flaviolin. Two cytochrome P450s (CYPs), StP450-1 and StP450-2, are present in the gene cluster. StTS was overexpressed in Escherichia coli BL21(DE3) and identified as a THN synthase. The synthesized THN can be easily oxidized into flaviolin by air. Both CYPs were reconstituted in E. coli BL21(DE3) and can oxidize flaviolin to form oligomers. The k(cat)/K(m) values for StP450-1 and StP450-2 were 0.28 and 0.71 min⁻¹ mM⁻¹, respectively. UV irradiation test showed that expression of StTS in E. coli BL21(DE3) significantly protects the cells from UV radiation, and coexpression of StTS and StP450-1 provides even stronger protection.
authors.affiliation: Department of Biological Engineering, Utah State University, 4105 Old Main Hill, Logan, UT 84322-4105, USA.
authors.forename: Jia
authors.initials: J
authors.lastname: Zeng
authors.forename: Richard
authors.initials: R
authors.lastname: Decker
authors.forename: Jixun
authors.initials: J
authors.lastname: Zhan
mesh_terms: D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000528:radiation effects; D015870:Gene Expression; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D009284:Naphthols / Q000378:metabolism; D009285:Naphthoquinones / Q000378:metabolism; D010084:Oxidation-Reduction; D010957:Plasmids; D011837:Radiation-Protective Agents / Q000378:metabolism; D016415:Sequence Alignment; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics; D013305:Streptomyces griseus / Q000201:enzymology / Q000235:genetics; D014466:Ultraviolet Rays
chemical_list: C121288:1,3,6,8-tetrahydroxynaphthalene; D001426:Bacterial Proteins; D007527:Isoenzymes; D009284:Naphthols; D009285:Naphthoquinones; D011837:Radiation-Protective Agents; C041659:flaviolin; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D000217:Acyltransferases; C020141:flavanone synthetase