Glycosylation of acyl carrier protein-bound polyketides during pactamycin biosynthesis

Abstract

Glycosylation is a common modification reaction in natural products biosynthesis and has been known to be a post assembly line tailoring process in glycosylated polyketide biosynthesis. Here, we show that in pactamycin biosynthesis glycosylation can take place on an acyl carrier protein (ACP)-bound polyketide intermediate. Using in vivo gene inactivation, chemical complementation, and in vitro pathway reconstitution we demonstrate that the 3aminoacetophenone moiety of pactamycin is derived from 3-aminobenzoic acid by a set of discrete polyketide synthase proteins via a 3-[3-aminophenyl]3-oxopropionyl-ACP intermediate. This ACP-bound intermediate is then glycosylated by an N-glycosyltransferase, PtmJ, providing a sugar precursor for the formation of the aminocyclopentitol core structure of pactamycin. This is the first example of glycosylation of a small molecule while tethered to a carrier protein.Additionally, we demonstrate that PtmO is a hydrolase that is responsible for the release of the ACP-bound product to a free β-ketoacid that subsequently undergoes decarboxylation.Glycosylation is one of the most ubiquitous and important transformations in nature and plays a central role in the structural and physiological aspects of living organisms. Glycosyltransferases are the family of enzymes that catalyze glycosylation, resulting in sugar-containing products. In natural products biosynthesis, glycosylation is generally considered to be a tailoring process that takes place later in the pathway after the backbone structure is formed. Some exceptions apply to certain natural products when glycosylation is directly involved in the formation of the core structure. Such phenomenon has been proposed to occur in the biosynthesis of at least two highly important microbial natural products, mitomycin and pactamycin. [1][2][3]

Identification and Biological Activity of NFAT-133 Congeners from Streptomyces pactum

“…S. pactum ATCC 27456 has been used as a model organism for the biosynthetic studies of the antitumor antibiotic pactamycin. ,− During the course of those studies, the potential of the strain to produce other bioactive natural products was recognized. Furthermore, inactivation of the NFAT-133 biosynthetic gene cluster in S. pactum ATCC 27456 Δ ptmTDQ , which does not produce pactamycin, ,, resulted in a mutant that also lacks the ability to produce NFAT-133 and a number of unknown minor metabolites .…”

Biosynthesis of cyclitols

“…159 The nitrogen atom at C-3 is derived from 3-aminobenzoic acid through N-glycosylation. 89,[160][161][162] The remaining nitrogen atom at C-1 is introduced aer N-glycosylation and dehydrogenation at the corresponding position by aminotransferase. 163 Therefore, the origin of the hydroxymethyl-containing cyclitols should be glucosamine-derived sugars, whose C-5 carbon atom is linked to the anomeric C-1 carbon atom of N-glycoside intermediate (Fig.…”

The chemistry and biology of natural ribomimetics and related compounds

“…27B ). 147 This highly unusual phenomenon, catalysed by the glycosyltransferase PtmJ, indicates that the formation of the C -ribomimetic unit takes place after the sugar precursor is attached to the 3-aminobenzoate/polyketide-derived intermediate ( e.g. , 3-aminoacetophenone).…”