Identification of an ATP-driven, osmoregulated glycine betaine transport system in Listeria monocytogenes

The ability of the gram-positive, food-borne pathogen Listeria monocytogene s to tolerate environments of elevated osmolarity and reduced temperature i s due in part to the transport and accumulation of the osmolyte glycine bet aine. Previously we showed that glycine betaine transport was: the result o f Na+-glycine betaine symport. In this report, we identify a second glycine betaine transporter from L. monocytogenes which is osmotically activated b ut does not require a high concentration of Na+ for activity. By using a po ol of Tn917-LTV3 mutants, a salt- and chill-sensitive mutant which was also found to be impaired in its ability to transport glycine betaine was isola ted. DNA sequence analysis of the region flanking the site of transposon in sertion revealed three open reading frames homologous to opuA hom Bacillus subtilis and proU from Escherichia coli, both of which encode glycine betai ne transport systems that belong to the superfamily of ATP-dependent transp orters. The three open reading frames are closely spaced, suggesting that t hey are arranged in an operon. Moreover, a region upstream from the first r eading frame was found to be homologous to the promoter regions of both opu A and proU. One unusual feature not shared with these other two systems is that the start codons for two of the open reading frames in L.. monocytogen es appear;to be TTG. That glycine:betaine uptake is nearly eliminated in th e mutant strain when it is assayed in the absence of Na+ is an indication t hat only the ATP-dependent transporter and the Na+-glycine betaine symporte r occur in L. monocytogenes.