Discrete acyltransferases associated with type I polyketide synthases and methods of use
View Patent ↗Genetic and biochemical characterization of the leinamycin biosynthesis gene cluster from Streptomyces atroolivaceus S-140 revealed two PKS genes, lnmI and lnmJ, that encode six PKS modules, none of which contains a cognate AT domain. The AT activity is provided in trans by a discrete protein, LnmG, which loads the malonyl coenzyme A extender unit onto the ACP domains of all six PKS modules. This finding provides a basis for methods of engineering modular polyketide synthases and polyketide synthase/nonribosomal peptide synthetases.
1. A method of loading an extender molecule on an acyl carrier protein (ACP) domain LnmJ of SEQ ID NO: 46 comprising the step of contacting said ACP domain LnmJ of SEQ ID NO: 46, with the extender molecule and a recombinantly expressed, discrete acyl transferase (AT) LnmG having the amino acid sequence of SEQ ID NO: 43, whereby the extender molecule is loaded onto the ACP domain LnmJ of SEQ ID NO: 46 by the catalytic activity of the discrete AT LnmG of SEQ ID NO. 43.
2. A method according to claim 1 wherein the method is carried out in vitro.
3. A method according to claim 1 wherein the method is carried out in a host cell.
4. A method according to claim 3 wherein the host cell includes a vector comprising a nucleic acid encoding the discrete AT LmnG of SEQ ID) NO. 43.
5. A method according to claim 4 wherein the host cell includes a different vector comprising a nucleic acid encoding the ACP domain LnmJ of SEQ ID NO 46.
6. A method according to claim 3 wherein the host cell is a bacterium.
7. A method according to claim 1 wherein the extender molecule is malonyl CoA.