Parsonage D , Newton GL , Holder RC , Wallace BD , Paige CL , Hamilton CJ , Dos Santos PC , Redinbo MR , Reid SD , Claiborne A
Biochemistry.2010 Aug 27 ; ():.
PMID: 20799687[PubMed - as supplied by publisher]
Bacillithiol (Cys-GlcN-malate, BSH) has recently been identified as a novel low-molecular-weight thiol in <i>Bacillus anthracis</i>, <i>Staphylococcus aureus</i>, and several other Gram-positive bacteria lacking glutathione and mycothiol. We have now characterized the first two enzymes for the BSH biosynthetic pathway in <i>B. anthracis</i>, which combine to produce alpha-D-glucosaminyl L-malate (GlcN-malate) from UDP-GlcNAc and L-malate. The structure of the GlcNAc-malate intermediate has been determined, as have the kinetic parameters for the <i>Ba</i>BshA glycosyltransferase (GlcNAc-malate) and the <i>Ba</i>BshB deacetylase (GlcN-malate). BSH is one of only two natural products reported to contain a malyl glycoside, and the crystal structure of the <i>Ba</i>BshA-UDP-malate ternary complex, determined in this work at 3.3 A resolution, identifies several active-site interactions important for the specific recognition of L-malate, but not other alpha-hydroxyacids, as acceptor substrate. In sharp contrast to the structures reported for the GlcNAc-1-D-<i>myo</i>-inositol-3-phosphate synthase (MshA) apo and ternary complex forms, there is no major conformational change observed in the structures of the corresponding <i>Ba</i>BshA forms. A mutant strain of <i>B. anthracis</i> deficient in the BshA glycosyltransferase fails to produce BSH, as predicted. This <i>B. anthracis bshA</i> locus (BA1558) has been identified in a transposon site hybridization study as required for growth, sporulation, or germination [Day, W.A., Jr., Rasmussen, S.L., Carpenter, B.M., Peterson, S.N., and Friedlander, A.M. (2007) <i>J. Bacteriol. 189</i>, 3296-3301], suggesting that the biosynthesis of BSH could represent a target for development of novel antimicrobials with broad spectrum activity against Gram-positive pathogens like <i>B. anthracis</i>. The metabolites that function in thiol redox buffering and homeostasis in <i>Bacillus</i> are not well understood, and we present a composite picture based on this and other recent work.