title: Biochemical and structural characterization of germicidin synthase: analysis of a type III polyketide synthase that employs acyl-ACP as a starter unit donor.
abstract: Germicidin synthase (Gcs) from Streptomyces coelicolor is a type III polyketide synthase (PKS) with broad substrate flexibility for acyl groups linked through a thioester bond to either coenzyme A (CoA) or acyl carrier protein (ACP). Germicidin synthesis was reconstituted in vitro by coupling Gcs with fatty acid biosynthesis. Since Gcs has broad substrate flexibility, we directly compared the kinetic properties of Gcs with both acyl-ACP and acyl-CoA. The catalytic efficiency of Gcs for acyl-ACP was 10-fold higher than for acyl-CoA, suggesting a strong preference toward carrier protein starter unit transfer. The 2.9 Å germicidin synthase crystal structure revealed canonical type III PKS architecture along with an unusual helical bundle of unknown function that appears to extend the dimerization interface. A pair of arginine residues adjacent to the active site affect catalytic activity but not ACP binding. This investigation provides new and surprising information about the interactions between type III PKSs and ACPs that will facilitate the construction of engineered systems for production of novel polyketides.
authors.affiliation: Life Sciences Institute, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Chemler
authors.forename: Tonia J
authors.initials: TJ
authors.lastname: Buchholz
authors.forename: Todd W
authors.initials: TW
authors.lastname: Geders
authors.forename: David L
authors.initials: DL
authors.lastname: Akey
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Rath
authors.forename: George E
authors.initials: GE
authors.lastname: Chlipala
authors.forename: Janet L
authors.initials: JL
authors.lastname: Smith
authors.forename: David H
authors.initials: DH
authors.lastname: Sherman
mesh_terms: D000213:Acyl Carrier Protein / Q000378:metabolism*; D000214:Acyl Coenzyme A / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D009154:Mutation; D048630:Polyketide Synthases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011753:Pyrones / Q000378:metabolism*; D015398:Signal Transduction; D048372:Streptomyces coelicolor / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D000213:Acyl Carrier Protein; D000214:Acyl Coenzyme A; D001426:Bacterial Proteins; D011753:Pyrones; C082790:germicidin; D048630:Polyketide Synthases