The r m s difference between the Cα atoms of the two monomers af

The r.m.s. difference between the Cα atoms of the two monomers after superposition is 0.38 Å, and the average B-factors of monomers A and B are 38.4 and 46.9 Å2, respectively. As with other alanine racemases, the AlrSP homodimer contains two active sites, each composed of residues from the

α/β barrel of one monomer and residues from the β-strand domain of the other. The pyridoxal phosphate Selleck GSK1838705A (PLP) cofactor is connected to Lys40 through an internal aldimine bond and resides inside the α/β barrel domain. Figure 1 Structure of alanine racemase from S. pneumoniae. (A) Ribbon diagram of the alanine racemase monomer with β-sheets colored green and α-helices colored gold. (B) Ribbon diagram of the alanine racemase dimer where one monomer is colored

blue and the opposite monomer red. The N’-pyridoxyl-lysine-5′-monophosphate or LLP residue (PLP cofactor covalently bound to lysine; black or grey spheres) resides in the α/β barrel domain of the active site. The active site is composed of residues from the α/β barrel domain of one monomer and residues from the β-strand domain of the other monomer. As an incidental finding, the AlrSP structure contained additional electron density within the A monomer, at the end of helix 1 in the N-terminal α/β barrel domain. This planar density CCI-779 nmr resembled a carboxylated aromatic ring, therefore a benzoic acid molecule, which fitted and refined well, was modeled into this region, even though the compound was not added to purification or crystallization conditions (topology and parameters obtained from the Hetero-compound Information Centre-Uppsala, HIC-UP [46]). It is situated some distance away from both the active site entryway and the dimer interface. Structural and biochemical comparison with closely related alanine racemases As noted in our previous publication [21], AlrSP displays a high level of sequence similarity with other alanine racemases. The structure-based sequence alignment in Figure 2 demonstrates this similarity

with alanine racemases from other Gram-positive bacteria: AlrEF (which has 52% sequence identity with AlrSP), AlrGS (46% identity), AlrBA (38% identity), and AlrSL (36% identity). Regions absolutely conserved across all of these enzymes include Farnesyltransferase the characteristic PLP binding site motif near the N-terminus (AVVKANAYGHG), the two catalytic amino acid residues of the active center (Lys40, Tyr263′; throughout this paper, primed numbers denote residues from the second monomer) and the eight residues making up the entryway to the active site (inner layer: Tyr263′, Tyr352, Tyr282′, and Ala169; middle layer: Arg307′, Ile350, Arg288′, and Asp170). Figure 2 Structure-based sequence alignment of the five solved alanine racemase structures from Gram-positive bacteria. Structures are from S. pneumoniae, G. stearothermophilus [29], E. faecalis [38], B.

Comments are closed.