The lytic polysaccharide monooxygenase of Pseudomonas aeruginosa promotes bacterial survival during systemic infection

ABSTRACT: Lytic polysaccharide monooxygenases (LPMOs) are a recently discovered enzyme family that cleave polysaccharides by oxidation. Despite proposed roles in bacterial virulence, no direct functional data exist to validate these claims. Here we show that CpbD, the LPMO of the opportunistic pathogen Pseudomonas aeruginosa (PA) is a virulence factor that promotes survival of the bacterium in whole human blood. CbpD was also shown to cleave the glycosidic bonds of the model substrate by an oxidative reaction, a feature that promoted by azurin and pyocyanin. Two redox-active virulence factors co-secreted with the LPMO. Combination of homology modelling, molecular dynamics simulations and small angle X-ray scattering demonstrated that CbpD is a monomeric tri-modular enzyme with highly flexible domain linkers, where domain positioning may be influenced by post translational modifications. Deletion of cbpD rendered P. aeruginosa unable to establish a lethal systemic infection and enhanced bacterial clearance in vivo. Improved bacterial survival of the wildtype was not attributable to dampening of pro-inflammatory responses by the protein, either ex vivo or in vivo. Further quantification of complement products in whole human blood revealed that CbpD reduced the performance of the terminal complement cascade. Importantly, CbpD inactivation by active site mutations abolished both enzyme activity in vitro and function ex vivo, indicating that ligand oxidation is crucial for CbpD virulence function. Finally, profiling of the bacterial and splenic proteome showed that the lack of this single enzyme resulted in substantial re-organization of the bacterial defense systems in vitro and the targeting of distinct host proteins/pathways in vivo.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human) Pseudomonas Aeruginosa Mus Musculus (mouse)

TISSUE(S): Spleen

SUBMITTER: Magnus Arntzen  

LAB HEAD: Gustav Vaaje-Kolstad

PROVIDER: PXD017971 | Pride | 2021-01-28

REPOSITORIES: Pride