title: Anatomy of the β-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate.
abstract: Alkyl branching at the β position of a polyketide intermediate is an important variation on canonical polyketide natural product biosynthesis. The branching enzyme, 3-hydroxy-3-methylglutaryl synthase (HMGS), catalyzes the aldol addition of an acyl donor to a β-keto-polyketide intermediate acceptor. HMGS is highly selective for two specialized acyl carrier proteins (ACPs) that deliver the donor and acceptor substrates. The HMGS from the curacin A biosynthetic pathway (CurD) was examined to establish the basis for ACP selectivity. The donor ACP (CurB) had high affinity for the enzyme (Kd = 0.5 μM) and could not be substituted by the acceptor ACP. High-resolution crystal structures of HMGS alone and in complex with its donor ACP reveal a tight interaction that depends on exquisite surface shape and charge complementarity between the proteins. Selectivity is explained by HMGS binding to an unusual surface cleft on the donor ACP, in a manner that would exclude the acceptor ACP. Within the active site, HMGS discriminates between pre- and postreaction states of the donor ACP. The free phosphopantetheine (Ppant) cofactor of ACP occupies a conserved pocket that excludes the acetyl-Ppant substrate. In comparison with HMG-CoA (CoA) synthase, the homologous enzyme from primary metabolism, HMGS has several differences at the active site entrance, including a flexible-loop insertion, which may account for the specificity of one enzyme for substrates delivered by ACP and the other by CoA.
authors: Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109; Chemical Biology Doctoral Program, University of Michigan, Ann Arbor, MI 48109;
authors: Finn P
authors: FP
authors: Maloney
authors: Center for Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography, University of California, San Diego, La Jolla, CA 92093;
authors: Lena
authors: L
authors: Gerwick
authors: Center for Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography, University of California, San Diego, La Jolla, CA 92093; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, CA 92093;
authors: William H
authors: WH
authors: Gerwick
authors: Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109; Department of Medicinal Chemistry, University of Michigan, Ann Arbor, MI 48109; Department of Chemistry, University of Michigan, Ann Arbor, MI 48109; Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI 48109;
authors: David H
authors: DH
authors: Sherman
authors: Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109; Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109 JanetSmith@umich.edu.
authors: Janet L
authors: JL
authors: Smith
mesh_terms: D000213:Acyl Carrier Protein / Q000737:chemistry* / Q000235:genetics; D000214:Acyl Coenzyme A / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D003521:Cyclopropanes / Q000737:chemistry; D006904:Hydroxymethylglutaryl-CoA Synthase / Q000737:chemistry* / Q000235:genetics; D048630:Polyketide Synthases / Q000737:chemistry* / Q000235:genetics; D061065:Polyketides / Q000737:chemistry*; D013302:Streptomyces / Q000235:genetics; D013379:Substrate Specificity; D013844:Thiazoles / Q000737:chemistry
chemical_list: D000213:Acyl Carrier Protein; D000214:Acyl Coenzyme A; D003521:Cyclopropanes; D061065:Polyketides; D013844:Thiazoles; C095843:curacin A; C008047:3-hydroxy-3-methylglutaryl-coenzyme A; D048630:Polyketide Synthases; D006904:Hydroxymethylglutaryl-CoA Synthase
keywords: HMG synthase; acyl carrier protein; curacin; natural products; polyketide synthase