Effect of superoxide dismutase and manganese on superoxide tolerance in Lactobacillus casei strain Shirota and analysis of multiple manganese transporters

The Lactobacillus casei/paracasei group accumulates a high level of manganese, which works to scavenge superoxide anions produced during aerobic growth. The genome of L. casei strain Shirota, however, also codes the gene for superoxide dismutase (SOD), sodA, which catalyzes the dismutation of superoxide anion into hydrogen peroxide and oxygen. We anticipated that the SOD and/or manganese may contribute to the aerobic growth of L. casei Shirota and tried to clarify how L. casei Shirota can eliminate the toxicity of superoxide anion. When the sodA of L. casei Shirota was cloned and expressed in Escherichia coli as well as in L. casei Shirota, there was no increase in SOD activity detected, meaning that the protein is in an inactive form, even if it is produced in L. casei Shirota. We next focused on the role of the manganese transport system of L. casei Shirota. One ABC-type manganese transporter (mtsCBA cluster) and three NRAMP-type manganese transporters (mntH1, mntH2, and mntH3) are coded in the genome. To clarify the role of these genes, we disrupted one or more of these manganese transporter genes in different combinations and analyzed the intracellular manganese levels. As a result, we found that NRAMP-type manganese transporters coded by mntH1 and mntH2 and ABCtype manganese transporter coded by mtsCBA cluster are complementarily involved in the accumulation of intracellular manganese and are necessary for aerobic growth of L. casei Shirota. These results indicate that intracellular manganese accumulated by multiple complementary manganese transporters, but not SOD, plays a pivotal role in tolerance to superoxide in L. casei Shirota.