title: Reconstitution of the FK228 biosynthetic pathway reveals cross talk between modular polyketide synthases and fatty acid synthase.
abstract: Functional cross talk between fatty acid biosynthesis and secondary metabolism has been discovered in several cases in microorganisms; none of them, however, involves a modular biosynthetic enzyme. Previously, we reported a hybrid modular nonribosomal peptide synthetase (NRPS)-polyketide synthase (PKS) pathway for the biosynthesis of FK228 anticancer depsipeptide in Chromobacterium violaceum strain 968. This pathway contains two PKS modules on the DepBC enzymes that lack a functional acyltransferase (AT) domain, and no apparent AT-encoding gene exists within the gene cluster or its vicinity. We report here that, through reconstitution of the FK228 biosynthetic pathway in Escherichia coli cells, two essential genes, fabD1 and fabD2, both encoding a putative malonyl coenzyme A (CoA) acyltransferase component of the fatty acid synthase complex, are positively identified to be involved in FK228 biosynthesis. Either gene product appears sufficient to complement the AT-less PKS modules on DepBC for polyketide chain elongation. Concurrently, a gene (sfp) encoding a putative Sfp-type phosphopantetheinyltransferase was identified to be necessary for FK228 biosynthesis as well. Most interestingly, engineered E. coli strains carrying variable genetic components produced significant levels of FK228 under both aerobic and anaerobic cultivation conditions. Discovery of the trans complementation of modular PKSs by housekeeping ATs reveals natural product biosynthesis diversity. Moreover, demonstration of anaerobic production of FK228 by an engineered facultative bacterial strain validates our effort toward the engineering of novel tumor-targeting bioagents.
authors.affiliation: Department of Biological Sciences, University of Wisconsin-Milwaukee, P.O. Box 413, Milwaukee, WI 53201, USA.
authors.forename: Shane R
authors.initials: SR
authors.lastname: Wesener
authors.forename: Vishwakanth Y
authors.initials: VY
authors.lastname: Potharla
authors.forename: Yi-Qiang
authors.initials: YQ
authors.lastname: Cheng
mesh_terms: D051080:Acyl-Carrier Protein S-Malonyltransferase / Q000235:genetics / Q000378:metabolism; D000693:Anaerobiosis; D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis; D001426:Bacterial Proteins / Q000378:metabolism; D053898:Biosynthetic Pathways; D002861:Chromobacterium / Q000235:genetics* / Q000378:metabolism*; D047630:Depsipeptides / Q000096:biosynthesis*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics; D054889:Fatty Acid Synthase, Type II / Q000235:genetics; D064429:Fatty Acid Synthases / Q000378:metabolism*; D005285:Fermentation; D015870:Gene Expression; D005818:Genetic Engineering; D008969:Molecular Sequence Data; D005810:Multigene Family; D046908:Peptide Biosynthesis, Nucleic Acid-Independent; D048630:Polyketide Synthases / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D017384:Sequence Deletion; D017855:Transferases (Other Substituted Phosphate Groups) / Q000378:metabolism
chemical_list: D000903:Antibiotics, Antineoplastic; D001426:Bacterial Proteins; D047630:Depsipeptides; D029968:Escherichia coli Proteins; C082662:phosphopantetheinyl transferase; D048630:Polyketide Synthases; C087123:romidepsin; D051080:Acyl-Carrier Protein S-Malonyltransferase; C494968:fabD protein, E coli; D064429:Fatty Acid Synthases; D017855:Transferases (Other Substituted Phosphate Groups); D054889:Fatty Acid Synthase, Type II