Interplay among Membrane-Bound Lytic Transglycosylase D1, the CreBC Two-Component Regulatory System, the AmpNG-AmpDI-NagZ-AmpR Regulatory Circuit, and L1/L2 β-Lactamase Expression in Stenotrophomonas maltophilia

Abstract

ABSTRACTLytic transglycosylases (LTs) are an important class of enzymes involved in peptidoglycan (PG) cleavage, with the concomitant formation of an intramolecular 1,6-anhydromuramoyl reaction product. There are six annotated LT genes in theStenotrophomonas maltophiliagenome, including genes for five membrane-bound LTs (mltA,mltB1,mltB2,mltD1, andmltD2) and a gene for soluble LT (slt). Six LTs ofS. maltophiliaKJ were systematically mutated, yielding the ΔmltA, ΔmltB1, ΔmltB2, ΔmltD1, ΔmltD2, and Δsltmutants. Inactivation ofmltD1conferred a phenotype of elevated uninduced β-lactamase activity. The underlying mechanism responsible for this phenotype was elucidated by the construction of several mutants and determination of β-lactamase activity. The expression of the genes assayed was assessed by quantitative reverse transcriptase PCR and a promoter transcription fusion assay. The results demonstrate that ΔmltD1mutant-mediated L1/L2 β-lactamase expression involved thecreBCtwo-component regulatory system (TCS) and theampNG-ampDI-nagZ-ampRregulatory circuit. The inactivation ofmltD1resulted inmltB1andmltD2upexpression in acreBC- andampNG-dependent manner. The overexpressed MltB1 and MltD2 activity contributed to the expression of the L1/L2 β-lactamase genes via theampNG-ampDI-nagZ-ampRregulatory circuit. These findings reveal, for the first time, a linkage between LTs, the CreBC TCS, theampNG-ampDI-nagZ-ampRregulatory circuit, and L1/L2 β-lactamase expression inS. maltophilia.