Crystal Structure and Enantiomer Selection by d-Alanyl Carrier Protein Ligase DltA from Bacillus cereus

Abstract

Ubiquitous D-alanylation of lipoteichoic acids modulates the surface charge and ligand binding of the gram-positive cell wall. Disruption of the bacterial DltABCD gene involved in teichoic acid alanylation, as well as inhibition of the DltA protein, has been shown to increase a gram-positive bacterium's susceptibility to antibiotics. The DltA D-alanyl carrier protein ligase promotes a two-step process starting with adenylation of D-alanine. We have determined the 2.0 A resolution crystal structure of a DltA protein from Bacillus cereus in complex with the D-alanine adenylate intermediate of the first reaction. Despite the low level of sequence similarity, the DltA structure resembles known structures of adenylation domains such as the acetyl-CoA synthetase. The enantiomer selection appears to be enhanced by the medium-sized side chain of Cys-269. The Ala-269 mutant protein shows marked loss of such selection. The network of noncovalent interactions between the D-alanine adenylate and DltA provides structure-based rationale for aiding the design of tight-binding DltA inhibitors for combating infectious gram-positive bacteria such as the notorious methicillin-resistant Staphylococcus aureus.

The wall teichoic acid and lipoteichoic acid polymers of Staphylococcus aureus

“…Recently, proteins involved in Rbo-P biosynthesis and incorporation have attracted particular interest, since WTA with Rbo-P residues are found in several human pathogens such as Staphylococcus saprophyticus (Schumacher-Perdreau et al, 1978), L. monocytogenes (Uchikawa et al, 1986), and Streptococcus pneumoniae (Fischer et al, 1993;Baur et al, 2009) in addition to S. aureus. Recently described crystal structures of proteins such as TagD (Fong et al, 2006), TarI (Baur et al, 2009), LtaS (Schirner et al, 2009;Lu et al, 2009), DltA (Du et al, 2008;Osman et al, 2009), and the Bacillus anthracis MnaA homolog (Velloso et al, 2008) represent the basis for rational drug design. Accordingly, specific inhibitors of DltA have recently been shown to render bacteria highly susceptible to CAMPs and cationic antibiotics (May et al, 2005) and to be very effective in clearing Gram-positive infections in vivo (Escaich et al, 2007).…”

Defining a Structural and Kinetic Rationale for Paralogous Copies of Phenylacetate-CoA Ligases from the Cystic Fibrosis Pathogen Burkholderia cenocepacia J2315

“…2b). Reminiscent of a leucine zipper, six leucine residues participate in hydrophobic interactions, with additional contributions from Tyr 30 , Phe 44 , and Phe 63 (Fig. 2b).…”

“…In the case of an AFE, the first two stages can be defined as the pre-and postadenylation state. It is noteworthy that capturing these different states in the context of one enzyme is rare, with DltA from Bacillus cereus serving as the only reported example (15,30). To explore the reorganization of key structural features in PaaK1 as the enzyme progresses through the adenylation reaction, we report the high resolution structures of PaaK1 in complex with ATP (Fig.…”

Structural Basis for the ATP-dependent Configuration of Adenylation Active Site in Bacillus subtilis o-Succinylbenzoyl-CoA Synthetase

“…At present, both the adenylate-forming and thioesterforming conformations have been structurally characterized for several ANL family members, including DltA from Bacillus subtilis (26,27), 4CBL (28,29), human medium-chain acyl-CoA synthetase 2A (ACSM2A) (30), and firefly luciferase (16,31).…”