Remodeling of Lipid A in Pseudomonas syringae pv. phaseolicola In Vitro

Abstract

<i>Pseudomonas</i> species infect a variety of organisms, including mammals and plants. Mammalian pathogens of the Pseudomonas family modify their lipid A during host entry to evade immune responses and to create an effective barrier against different environments, for example by removal of primary acyl chains, addition of phosphoethanolamine (<i>P</i>-EtN) to primary phosphates, and hydroxylation of secondary acyl chains. For <i>Pseudomonas syringae</i> pv. <i>phaseolicola</i> (<i>Pph</i>) 1448A, an economically important pathogen of beans, we observed similar lipid A modifications by mass spectrometric analysis. Therefore, we investigated predicted proteomes of various plant-associated <i>Pseudomonas</i> spp. for putative lipid A-modifying proteins using the well-studied mammalian pathogen <i>Pseudomonas aeruginosa</i> as a reference. We generated isogenic mutant strains of candidate genes and analyzed their lipid A. We show that the function of PagL, LpxO, and EptA is generally conserved in <i>Pph</i> 1448A. PagL-mediated de-acylation occurs at the distal glucosamine, whereas LpxO hydroxylates the secondary acyl chain on the distal glucosamine. The addition of <i>P</i>-EtN catalyzed by EptA occurs at both phosphates of lipid A. Our study characterizes lipid A modifications in vitro and provides a useful set of mutant strains relevant for further functional studies on lipid A modifications in <i>Pph</i> 1448A.