Abstact

Lytic transglycosylases (LTs) belong to a family of enzymes that turnover the bacterial cell-wall peptidoglycan through a nonhydrolytic cleavage of the beta(1-4) glycosidic bond, generating a hallmark 1,6-anhydromuramyl moiety in the reaction products. LTs are essential for numerous cellular processes, including cell-wall maturation, peptidoglycan recycling, cell division, and the assembly of multiprotein complexes. Their functional diversity underscores their biological significance. Family 3 LTs are distinguished by their EF-hand Ca(2+)-binding motif and are classified into two subfamilies. Subfamily 3B members, including Pseudomonas aeruginosa SltB2, possess a peptidoglycan-binding domain absent in subfamily 3A. In this study, we present the structural characterization of P. aeruginosa SltB2. The high-resolution crystal structure of SltB2 reveals a unique modular architecture shaped by the specific arrangement of its PG-binding domain and distinct differences in the organization of key residues surrounding the catalytic Glu residue compared to other family 3 members. A model of interaction between SltB2 and the peptidoglycan is proposed, which accounts for the enzyme's tolerance to peptide stems and reveals particular features at site +2, due to the unique arrangement of the PG-binding domain, explaining its preferred exolytic activity. Comparative structural analyses of Family 3 LTs provide insights into substrate recognition and enzymatic function, advancing our understanding of bacterial cell-wall remodeling mechanisms.