Rs. Gokhale et al., Mechanism and specificity of the terminal thioesterase domain from the erythromycin polyketide synthase, CHEM BIOL, 6(2), 1999, pp. 117-125
Background: Polyketides are important compounds with antibiotic and antican
cer activities. Several modular polyketide syntheses (PKSs) contain a termi
nal thioesterase (TE) domain probably responsible for the release and conco
mitant cyclization of the fully processed polyketide chain. Because the TE
domain influences qualitative aspects of product formation by engineered PK
Ss, its mechanism and specificity are of considerable interest.
Results: The TE domain of the 6-deoxyerythronolide B synthase was overexpre
ssed in Escherichia coli, When tested against a set of N-acetyl cysteamine
thioesters the TE domain did not act as a cyclase, but showed significant h
ydrolytic specificity towards substrates that mimic important features of i
ts natural substrate. Also the overall rate of polyketide chain release was
strongly enhanced by a covalent connection between the TE domain and the t
erminal PKS module (by as much as 100-fold compared with separate TE and PK
S 'domains'),
Conclusions: The inability of the TE domain alone to catalyze cyclization s
uggests that macrocycle formation results from the combined action of the T
E domain and a PKS module, The chain-length and stereochemical preferences
of the TE domain might be relevant in the design and engineered biosynthesi
s of certain novel polyketides. Our results also suggest that the TE domain
might loop back to catalyze the release of polyketide chains from both ter
minal and pre-terminal modules, which may explain the ability of certain na
turally occurring PKSs, such as the picromycin synthase, to generate both 1
2-membered and 14-membered macrolide antibiotics.