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  • Role of a SpoVA protein in dipicolinic acid uptake into developing spores of Bacillus subtilis.

Role of a SpoVA protein in dipicolinic acid uptake into developing spores of Bacillus subtilis.

Journal of bacteriology (2012-02-14)
Yunfeng Li, Andrew Davis, George Korza, Pengfei Zhang, Yong-qing Li, Barbara Setlow, Peter Setlow, Bing Hao
ABSTRACT

The proteins encoded by the spoVA operon, including SpoVAD, are essential for the uptake of the 1:1 chelate of pyridine-2,6-dicarboxylic acid (DPA(2,6)) and Ca(2+) into developing spores of the bacterium Bacillus subtilis. The crystal structure of B. subtilis SpoVAD has been determined recently, and a structural homology search revealed that SpoVAD shares significant structural similarity but not sequence homology to a group of enzymes that bind to and/or act on small aromatic molecules. We find that molecular docking placed DPA(2,6) exclusively in a highly conserved potential substrate-binding pocket in SpoVAD that is similar to that in the structurally homologous enzymes. We further demonstrate that SpoVAD binds both DPA(2,6) and Ca(2+)-DPA(2,6) with a similar affinity, while exhibiting markedly weaker binding to other DPA isomers. Importantly, mutations of conserved amino acid residues in the putative DPA(2,6)-binding pocket in SpoVAD essentially abolish its DPA(2,6)-binding capacity. Moreover, replacement of the wild-type spoVAD gene in B. subtilis with any of these spoVAD gene variants effectively eliminated DPA(2,6) uptake into developing spores in sporulation, although the variant proteins were still located in the spore inner membrane. Our results provide direct evidence that SpoVA proteins, in particular SpoVAD, are directly involved in DPA(2,6) movement into developing B. subtilis spores.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
2,6-Pyridinedicarboxylic acid, 99%
Supelco
2,6-Pyridinedicarboxylic acid, suitable for ion chromatography, ≥99.5% (T)
Supelco
2,6-Pyridinedicarboxylic acid concentrate, 0.02 M C7H5NO4 in water (0.04N), suitable for ion chromatography, eluent concentrate