Mutation ofmltGincreases peptidoglycan fragment release, cell size, and antibiotic susceptibility inNeisseria gonorrhoeae

Abstract

AbstractInfection with the Gram-negative speciesNeisseria gonorrhoeaeleads to inflammation that is responsible for the disease symptoms of gonococcal urethritis, cervicitis, and pelvic inflammatory disease. During growth these bacteria release significant amounts of peptidoglycan (PG) fragments which elicit inflammatory responses in the human host. To better understand the mechanisms involved in PG synthesis and breakdown inN. gonorrhoeae, we characterized the effects of mutation ofmltG. MltG has been identified in other bacterial species as a terminase that stops PG strand growth by cleaving the growing glycan. Mutation ofmltGinN. gonorrhoeaedid not affect bacterial growth rate but resulted in increased PG turnover, more cells of large size, decreased autolysis under non-growth conditions, and increased sensitivity to antibiotics that affect PG crosslinking. AnmltGmutant released greatly increased amounts of PG monomers, PG dimers, and larger oligomers. In themltGbackground, mutation of eitherltgAorltgD, encoding the lytic transglycosylases responsible for PG monomer liberation, resulted in wild-type levels of PG monomer release. Bacterial two-hybrid assays identified positive interactions of MltG with synthetic penicillin-binding proteins PBP1 and PBP2 and the PG-degrading endopeptidase PBP4 (PbpG). These data are consistent with MltG acting as a terminase inN. gonorrhoeaeand suggest that absence of MltG activity results in excessive PG growth and extra PG in the sacculus that must be degraded by lytic transglycosylases including LtgA and LtgD. Furthermore, absence of MltG causes a cell wall defect that is manifested as large cell size and antibiotic sensitivity.ImportanceNeisseria gonorrhoeaeis unusual in that the bacteria release larger amounts of cell wall material as they grow as compared to related bacteria, and the released cell wall fragments induce inflammation that leads to tissue damage in infected people. The study of MltG revealed the importance of this enzyme for controlling cell wall growth, cell wall fragment production, and bacterial cell size and suggest a role for MltG in a cell wall synthesis and degradation complex. The increased antibiotic sensitivities of anmltGmutant suggest that an antimicrobial drug inhibiting MltG would be useful in combination therapy to restore the sensitivity of the bacteria to cell wall targeting antibiotics to which the bacteria are currently resistant.