The phosphopantetheinyl transferase KirP activates the ACP and PCP domains of the kirromycin NRPS/PKS of Streptomyces collinus Tü 365

Abstract

The main steps in the biosynthesis of complex secondary metabolites such as the antibiotic kirromycin are catalyzed by modular polyketide synthases (PKS) and/or nonribosomal peptide synthetases (NRPS). During antibiotic assembly, the biosynthetic intermediates are attached to carrier protein domains of these megaenzymes via a phosphopantetheinyl arm. This functional group of the carrier proteins is attached post-translationally by a phosphopantetheinyl transferase (PPTase). No experimental evidence exists about how such an activation of the carrier proteins of the kirromycin PKS/NRPS is accomplished. Here we report on the characterization of the PPTase KirP, which is encoded by a gene located in the kirromycin biosynthetic gene cluster. An inactivation of the kirP gene resulted in a 90% decrease in kirromycin production, indicating a substantial role for KirP in the biosynthesis of the antibiotic. In enzymatic assays, KirP was able to activate both acyl carrier protein and petidyl carrier domains of the kirromycin PKS/NRPS. In addition to coenzyme A (CoA), which is the natural substrate of KirP, the enzyme was able to transfer acyl-phosphopantetheinyl groups to the apo forms of the carrier proteins. Thus, KirP is very flexible in terms of both CoA substrate and carrier protein specificity. Our results indicate that KirP is the main PPTases that activates the carrier proteins in kirromycin biosynthesis.

The Insect Pathogen Serratia marcescens Db10 Uses a Hybrid Non-Ribosomal Peptide Synthetase-Polyketide Synthase to Produce the Antibiotic Althiomycin

“…The gene encoding the PPTase required for activation of an NRPS or PKS enzyme may or may not be encoded in the same region of the genome as the NRPS or PKS biosynthetic gene cluster [30]. The alb1–alb6 gene cluster does not encode a PPTase enzyme; therefore we extended our search for the PPTase needed for althiomycin biosynthesis to the entire genome of S. marcescens Db10.…”

The phosphopantetheinyl transferases: catalysis of a post-translational modification crucial for life

“…52 Kirromycin was discovered in 1974 by Wolf et al, 133 and >200 publications have discussed its biosynthesis and mechanisms of action. In 2007, Weber et al characterized the PKS-NRPS hybrid in detail and identify the PPTase KirP, essential for phosphopantetheinylating the large number of acyl- and peptidyl carrier proteins.…”

“…134 Interestingly, inactivation of KirP does not result in the total abolishment of kirromycin biosynthesis. 52 …”

A Single Sfp-Type Phosphopantetheinyl Transferase Plays a Major Role in the Biosynthesis of PKS and NRPS Derived Metabolites in Streptomyces ambofaciens ATCC23877

“…In addition, SCO6673-like could be involved in the activation of the ACP and PCP domains encoded by the still cryptic type I PKS and NRPS gene clusters identified in the S. ambofaciens ATCC23877 chromosome by genome mining. Other members of the F/KES family in Streptomyces are also flexible in terms of carrier proteins such as KirP of Streptomyces collinus Tü365 which targets ACP and PCP domains of the kirromycin NRPS/PKS [47], but none of them have been reported to activate the two types of carrier proteins from different secondary metabolite biosynthetic pathways in vivo .…”