A specific role of the Saccharopolyspora erythraea thioesterase II gene in the function of modular polyketide synthases

Abstract

Bacterial modular polyketide synthase (PKS) genes are commonly associated with another gene that encodes a thioesterase II (TEII) believed to remove aberrantly loaded substrates from the PKS. Co-expression of the Saccharopolyspora erythraea ery-ORF5 TEII and eryA genes encoding 6-deoxyerythronolide B synthase (DEBS) in Streptomyces hosts eliminated or significantly lowered production of 8,89-deoxyoleandolide [15-nor-6-deoxyerythronolide B (15-nor-6dEB)], which arises from an acetate instead of a propionate starter unit. Disruption of the TEII gene in an industrial Sac. erythraea strain caused a notable amount of 15-norerythromycins to be produced by utilization of an acetate instead of a propionate starter unit and also resulted in moderately lowered production of erythromycin compared with the amount produced by the parental strain. A similar behaviour of the TEII gene was observed in Escherichia coli strains that produce 6dEB and 15-methyl-6dEB. Direct biochemical analysis showed that the ery-ORF5 TEII enzyme favours hydrolysis of acetyl groups bound to the loading acyl carrier protein domain (ACP L ) of DEBS. These results point to a clear role of the TEII enzyme, i.e. removal of a specific type of acyl group from the ACP L domain of the DEBS1 loading module.

Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate

“…Disruption of the TE II gene associated with 6-deoxyerythronolide B synthase (DEBS) of Saccharopolyspora erythraea resulted in a notable increase in shunt products formed by utilization of acetate instead of propionate as a starter unit, while the combined level of all polyketides produced was reduced by less than 20% (20). The DEBS TE II exhibited a strong in vitro preference for the acetyl group bound to the ACP from the loading module of DEBS.…”

“…These discrete, 25-to 29-kDa proteins were first identified in some mammalian fatty acid synthase (FAS) complexes, where they are alternative chain-terminating enzymes with affinities for medium-chain-length fatty acids (38). In PKS and NRPS systems they appear to have an "editing" role, removing nonreactive acyl residues blocking the multienzyme (16,36,50), removing amino acids that cannot be elongated (48), and/or controlling starter unit selection (20). However, recent reports on polyether ionophore antibiotic biosynthesis have shown that TE IIs are responsible for the release of this class of polyketides by hydrolysis (15,28,29).…”

“…The DEBS TE II exhibited a strong in vitro preference for the acetyl group bound to the ACP from the loading module of DEBS. Therefore, it was proposed that removal of acetate from the ACP of the loading module is the main function of this TE II (20). This paper deals with the TE II ScoT associated with the type I modular PKS Cpk from Streptomyces coelicolor A3(2) (32).…”

Structure and Functional Analysis of RifR, the Type II Thioesterase from the Rifamycin Biosynthetic Pathway

“…TEIIs are able to hydrolyze substrates attached to carrier domains from their native pathway as well as other pathways (6,20,28).…”

Identification of a Gene Cluster of Polyether Antibiotic Lasalocid fromStreptomyces lasaliensis

“…Type II TE serves a conventional editing function to remove aberrantly attached acyl intermediates from the PKS. 34,35) lsd9 encodes a putative crotonyl-CoA reductase, which is a key enzyme for providing ethylmalonylCoA, an essential extender unit of the lasalocid polyketide backbone, 36) and displays high homology to PlmT7 (Streptomyces sp. HK803, 80%).…”