The Mechanism of Dehydrating Bimodules in trans‐Acyltransferase Polyketide Biosynthesis: A Showcase Study on Hepatoprotective Hangtaimycin

Abstract

A bioassay‐guided fractionation led to the isolation of hangtaimycin (HTM) from Streptomyces spectabilis CCTCC M2017417 and the discovery of its hepatoprotective properties. Structure elucidation by NMR suggested the need for a structural revision. A putative HTM degradation product was also isolated and its structure was confirmed by total synthesis. The biosynthetic gene cluster was identified and resembles a hybrid trans‐AT PKS/NRPS biosynthetic machinery whose first PKS enzyme contains an internal dehydrating bimodule, which is usually found split in other trans‐AT PKSs. The mechanisms of such dehydrating bimodules have often been proposed, but have never been deeply investigated. Here we present in vivo mutations and in vitro enzymatic experiments that give first and detailed mechanistic insights into catalysis by dehydrating bimodules.

Structural characterization and hepatoprotective activity of an acidic polysaccharide from Ganoderma lucidum

“… Xiao et al, 2019 ). Stress, excessive diet, alcohol and drugs abuse can cause liver injure, which may further develop into chronic fatigue, coagulopathy, liver failure and even cancers ( Luo et al, 2021 ). Liver diseases which develop from liver injury have brought enormous burdens for society and economics.…”

Cytochrome P450 Monooxygenase for Catalyzing C-42 Hydroxylation of the Glycine-Derived Fragment in Hangtaimycin Biosynthesis

“…On the right end of the cluster downstream of the trans -AT gene htm10 , the htm11 gene encodes a cytochrome P450 monooxygenase. An essential role for HTM biosynthesis was verified for the htm10 homologous gene htmA7 in S. spectabilis CCTCC M2017417 . HTM production was abolished in the htm11 deletion mutant (Htm11KO), but TDD was detected at levels similar to the wild-type strain, indicating that an HTM biosynthetic intermediate bearing the TDD moiety was produced in the Htm11KO (Figures D and A).…”

“…The cluster encodes six NRPS modules, two in PKS/NRPS hybrid Htm2 and four in NRPS Htm4 and 5 (Figure A, Table S3). A ClusterBlast analysis against the antiSMASH database indicated that the htm cluster was present and conserved in some S. spectabilis strains, although the gene arrangement and size differed slightly from the htm cluster in S. spectabilis CCTCC M2017417 (Figure S1). To confirm the htm cluster and to identify its boundaries, six genes in or flanking the region of the cluster were inactivated in S. spectabilis CPCC 200148 by gene knockout (Figures S2–S7).…”

“…As opposed to canonical cis -AT PKSs, trans -AT PKSs lack the AT domain in each extension module but share a free-standing AT. , The htm10 gene in the htm cluster encodes a protein containing two AT domains, homologous to htmA7 in S. spectabilis CCTCC M2017417 (Figure S1). According to the biosynthetic paradigm of trans -AT PKSs, the acetyl-loading AT (GNAT) domain of the loading module in Htm1 decarboxylates malonyl-CoA to produce the acetyl-CoA starter followed by two rounds of chain extension and dehydration, as elucidated by Luo et al Htm2 is a hybrid trans -AT PKS/NRPS (modules 5–10), in which module 5 is predicted to assemble an alanine and module 8 is predicted to assemble a glycine (Figure ). Htm3 is a hybrid trans -AT PKS containing two tandem acceptor acyl carrier protein domains (ACP A , Figure S9C) where β-branching may take place.…”

“…Recently, Luo et al reported another HTM producer, S. spectabilis CCTCC M2017417, in screening compounds for hepatoprotective activity. 2 They identified the HTM biosynthetic gene cluster (BGC), proposed its biosynthesis process, and resolved the catalytic mechanism of its first polyketide synthase (PKS) enzyme. However, details of the biosynthetic mechanism of this unusual compound need to be further clarified.…”