LIPID A REGIOSELECTIVITY OF THE ESCHERICHIA COLI PALMITOYLTRANSFERASE PAGP

Abstract

The outer membrane palmitoyltransferase PagP possesses regioselectivity for the palmitoylation of the (R)-3-hydroxymyristate chain at position 2 on the proximal glucosamine unit of lipid A. The residues Arg45 and Arg49 in the L1 loop appear to poise their guanidinium groups so as to interact with the proximal and distal phosphate groups at positions 1 and 4’ of lipid A, respectively. Both single and double substitution of these arginine residues with serine has no effect on the folding, stability, phospholipase and palmitoyltransferase activities. Additionally, the arginine to serine substitutions display wild-type regioselectivity and specific activity in the palmitoylation of the biosynthetic precursor lipid IVA as indicated by collision induced fragmentation MS/MS. In vivo, lipid A analysis in a msbB/pagP deletion strain shows no difference in acylation pattern as compared to the wild-type. These results establish both in vitro and in vivo that the arginine to serine substitutions have no effect on lipid A regioselectivity.