Crystal structure and acyl chain selectivity of Francisella novicida LpxA, the first enzyme in lipid A biosynthesis

Abstract

The first step of lipid A biosynthesis in Escherichia coli is catalyzed by LpxA (EcLpxA), an acyltransferase selective for UDP‐GlcNAc and R‐3‐hydroxymyristoyl‐acyl carrier protein (ACP). While EcLpxA is selective for R‐3‐hydroxymyristoyl‐acyl chain (C14), Francisella novicida LpxA (FnLpxA) is thought to be selective for longer acyl chain (C18) in vivo. Here we report the 2.06 Å crystal structure of the FnLpxA. The N‐terminal parallel beta‐helix (LbetaH) and C‐terminal alpha‐helical domain are similar to the structures of LpxA's solved so far. The preference for the longer acyl chain is thought to be derived from the L180 ‐ G188 region which takes a wider turn with two extra residues compared to EcLpxA, providing a flexible pocket to accommodate longer acyl chain. While purified FnLpxA prefers R,S‐hydroxypalmitoyl‐ACP over R,S‐3‐hydroxymyristoyl‐ACP under standard in vitro assay conditions, complementation of an LpxA knockout Escherichia coli strain with wild‐type FnLpxA generates lipid A species mainly of the usual R‐3‐hydroxymyristate (C14) at positions 2 and 2′, with smaller population of one or two longer hydroxy fatty acids (C16), as judged by mass spectrometry. Our results indicate the acyl chain length of lipid A is determined in several levels including the acyl carrier protein pool, LpxA specificity, and beyond.