Structures of Bacillus subtilis PdaA, a family 4 carbohydrate esterase, and a complex with N‐acetyl‐glucosamine

Abstract

Family 4 carbohydrate esterases deacetylate polymeric carbohydrate substrates such as chitin, acetyl xylan and peptidoglycan. Although some of these enzymes have recently been enzymologically characterised, neither their structure nor their reaction mechanism has been defined. Sequence conservation in this family has pointed to a conserved core, termed the NodB homology domain. We describe the cloning, purification and 1.9 A crystal structure of PdaA, a peptidoglycan deacetylase from Bacillus subtilis. The enzyme assumes a fold related to a ðb=aÞ 8 barrel, with a long groove on the surface of the protein that harbours all conserved residues. A complex with the substrate analogue N-acetyl-glucosamine was refined to 2.25 A resolution, revealing interactions of an aspartic acid and three histidines, all conserved in the NodB homology domain, with the ligand. The PdaA structure provides a template for interpreting the wealth of sequence data on family 4 carbohydrate esterases in a structural context and represents a first step towards understanding the reaction mechanism of this family of enzymes.

Structure and metal-dependent mechanism of peptidoglycan deacetylase, a streptococcal virulence factor

“…So far, only one other CE-4 structure has been reported, that of BsPdaA (11). Strikingly, the equivalent of SpPgdA Asp-276, one of the zinc-binding ligands, is an asparagine in PdaA, and the backbone around this residue adopts a dramatically different conformation, with the side chain buried in the core of the protein (Fig.…”

“…Two of the conserved histidines come together in the bottom of the active site, yet soaking studies were inconclusive as to the possible role of these histidines as metal-binding residues. A complex with GlcNAc was also reported, although it was not clear whether this represented productive substrate binding (11). Thus, further structural and biochemical studies are required to define the mechanism of action of the CE-4 family.…”

“…The BsPdaA enzyme deacetylates peptidoglycan N-acetyl muramic acid residues, leading to the formation of Dlactam structures (12). The BsPdaA NodB homology domain appears to adopt an (␣͞␤) 8 fold, with a groove running over the surface of the protein harboring the majority of the conserved residues (11). Two of the conserved histidines come together in the bottom of the active site, yet soaking studies were inconclusive as to the possible role of these histidines as metal-binding residues.…”

“…Recently, the first structure of a CE-4 family member, the Bacillus subtilis peptidoglycan deacetylase (BsPdaA), has been reported (11). The BsPdaA enzyme deacetylates peptidoglycan N-acetyl muramic acid residues, leading to the formation of Dlactam structures (12).…”

Structure and Mechanism of Chitin Deacetylase from the Fungal Pathogen Colletotrichum lindemuthianum,

“…These studies have suggested that chitin is only partially deacetylated in a specific manner that may prevent recognition by either chitinases or chitosanases. The first structure of a CE-4 family member, B. subtilis PdaA, was reported recently (33). The protein adopts a ( /R) 8 fold with a putative substratebinding groove harboring the majority of the conserved residues.…”