Structure and Mechanism of Staphylococcus aureus TarS, the Wall Teichoic Acid β-glycosyltransferase Involved in Methicillin Resistance

Abstract

In recent years, there has been a growing interest in teichoic acids as targets for antibiotic drug design against major clinical pathogens such as Staphylococcus aureus, reflecting the disquieting increase in antibiotic resistance and the historical success of bacterial cell wall components as drug targets. It is now becoming clear that β-O-GlcNAcylation of S. aureus wall teichoic acids plays a major role in both pathogenicity and antibiotic resistance. Here we present the first structure of S. aureus TarS, the enzyme responsible for polyribitol phosphate β-O-GlcNAcylation. Using a divide and conquer strategy, we obtained crystal structures of various TarS constructs, mapping high resolution overlapping N-terminal and C-terminal structures onto a lower resolution full-length structure that resulted in a high resolution view of the entire enzyme. Using the N-terminal structure that encapsulates the catalytic domain, we furthermore captured several snapshots of TarS, including the native structure, the UDP-GlcNAc donor complex, and the UDP product complex. These structures along with structure-guided mutants allowed us to elucidate various catalytic features and identify key active site residues and catalytic loop rearrangements that provide a valuable platform for anti-MRSA drug design. We furthermore observed for the first time the presence of a trimerization domain composed of stacked carbohydrate binding modules, commonly observed in starch active enzymes, but adapted here for a poly sugar-phosphate glycosyltransferase.

Salt-Induced Stress Stimulates a Lipoteichoic Acid-Specific Three-Component Glycosylation System in Staphylococcus aureus

“…The functions of DltB and DltD are not as clear but are thought to transport the DltC-D-Ala intermediate across the membrane and subsequently D-alanylate TA acceptor substrates. Glycosylation of TAs has remained relatively underexamined by comparison, but recent studies have made progress in demonstrating the roles of TarM and TarS in transferring ␣and ␤-GlcNAc, respectively, to WTA in S. aureus (24)(25)(26)(27). WTA modification with ␤-GlcNAc imparts beta-lactam resistance, is critical to colonization, and plays a role alongside D-Ala modifications in cell division and morphogenesis (28,29).…”

Constructing and deconstructing the bacterial cell wall

“…Intimate structural synergy is now recognized between the peptidoglycan and the teichoic acids of the Gram-positive bacteria, 76 and between the peptidoglycan and the outer membrane of the Gramnegative bacteria. 77,78 Indeed, new opportunities for antibiotic discovery have been identified that interfere with teichoic acid integration into the cell wall of Gram-positive bacteria 69,[79][80][81][82][83][84] and with respect to lipopolysaccharide transport in Gram-negative bacteria. [85][86][87][88][89][90] Within this universe of opportunity, we exemplify our own efforts toward the mechanistic and structural understanding of key enzymes of the bacteria with roles in cell envelope biosynthesis and antibiotic resistance.…”

Characterization of Wall Teichoic Acid Degradation by the Bacteriophage ϕ29 Appendage Protein GP12 Using Synthetic Substrate Analogs

“…Moreover, the GP12 trimer has the capacity to accommodate multiple polymeric substrates (27), a property associated with processively acting enzymes (45). Even so, taking into account that GP12 catalyzes two distinct hydrolytic reactions, it would seem that non-processive polymer degradation is a likely mode of action for this enzyme.…”