The Retaining Pse5Ac7Ac Pseudaminyltransferase KpsS1 Defines a Previously Unreported glycosyltransferase family (GT118)

Cell surface sugar 5,7-diacetyl pseudaminic acid (Pse5Ac7Ac) is a bacterial analogue of the ubiquitous sialic acid, Neu5Ac, and contributes to the virulence of a number of multidrug resistant bacteria, including ESKAPE pathogens Pseudomonas aeruginosa, and Acinetobacter baumannii. Despite its discovery in the surface glycans of bacteria over thirty years ago, to date no glycosyltransferase enzymes (GTs) dedicated to the synthesis of a pseudaminic acid glycosidic linkage have been unequivocally characterised in vitro. Herein we demonstrate that A. baumannii KpsS1 is a dedicated pseudaminyltransferase enzyme (PseT) which constructs a Pse5Ac7Ac-alpha(2,6)-Glcp linkage, and proceeds with retention of anomeric configuration. We utilise this PseT activity in tandem with the biosynthetic enzymes required for CMP-Pse5Ac7Ac assembly, in a two-pot, seven enzyme synthesis of an alpha-linked Pse5Ac7Ac glycoside. Due to its unique activity and protein sequence, we also assign KpsS1 as the prototypical member of a previously unreported GT family (GT118). Pse5Ac7Ac glycosyltransferase (GT) activity has been demonstrated in characterisation of the retaining pseudaminyltransferase (PseT) KpsS1, the prototypical member of a previously unreported GT family (GT118). KpsS1 is utilised in a two-pot, seven enzyme synthesis of a glycoside terminating in Pse5Ac7Ac, an important bacterial analogue of the ubiquitous sialic acid Neu5Ac.+ image