The role of salicylic acid in iron metabolism was examined in two wild-type strains (mc 2 155 and NCIMB 8548) and three mutant strains (mc 2 1292 [lacking exochelin], SM3 [lacking iron-dependent repressor protein IdeR] and S99 [a salicylate-requiring auxotroph derived in this study]) of Mycobacterium smegmatis. Synthesis of salicylate in SM3 was derepressed even in the presence of iron, as was synthesis of the siderophores exochelin, mycobactin, and carboxymycobactin. S99 was dependent on salicylate for growth and failed to grow with the three ferrisiderophores, suggesting that salicylate fulfills an additional function(s) other than being a precursor of mycobactin and carboxymycobactin. Salicylic acid at 100 g/ml repressed the formation of a 29-kDa cell envelope protein (putative exochelin receptor protein) in S99 grown both iron deficiently and iron sufficiently. In contrast, synthesis of this protein was affected only under iron-limited conditions in the parent strain, mc 2 155, and remained unaltered in SM3, suggesting an interaction between the IdeR protein and salicylate. Thus, salicylate may also function as a signal molecule for recognition of cellular iron status. Growth of all strains and mutants with p-aminosalicylate (PAS) at 100 g/ml increased salicylate accumulation between three-and eightfold under both iron-limited and iron-sufficient growth conditions and decreased mycobactin accumulation by 40 to 80% but increased carboxymycobactin accumulation by 50 to 55%. Thus, although PAS inhibited salicylate conversion to mycobactin, presumptively by blocking salicylate AMP kinase, PAS also interferes with the additional functions of salicylate, as its effect was heightened in S99 when the salicylate concentration was minimal.Mycobacterium smegmatis, a saprophytic mycobacterium, secretes two extracellular siderophores (exochelin and carboxymycobactin) and has an intracellular siderophore termed mycobactin (28, 30). De Voss et al. (8) demonstrated the importance of the siderophores mycobactin and carboxymycobactin in M. tuberculosis by showing that a mutant incapable of producing these two siderophores failed to grow within macrophages. Carboxymycobactin is structurally related to mycobactin but is probably not derived from it (2). While carboxymycobactin is the sole extracellular siderophore of pathogenic mycobacteria (30), it is only a minor one in saprophytic mycobacteria, which use exochelin as the major siderophore for iron acquisition (28). Salicylic acid is produced extracellularly by most mycobacteria in greatly increased quantities (up to 17 g/ml) when grown under iron-deficient conditions (28, 33) as opposed to iron-sufficient conditions. Although salicylate is a precursor of mycobactin (31), mycobactin could not spare the requirement for salicylate in a salicylate-requiring auxotrophic mutant of M. smegmatis (32), suggesting that salicylate fulfills a second but unknown role in this bacterium.Salicylate and its derivatives are important as analgesics, antipyretics, anticoagulants, and anti-infl...

Three-point transduction analysis was employed to determine the sequence of four structural and two regulatory methionine genes (metA, B, F, HandmetI, J, respectively) on two apparently adjacent transduction fragments inSalmonella typhimurium. These fragments were subsequently orientated with respect to each other and the rest of the linkage map by the analysis of recombinants obtained in conjugation experiments. The following clockwise sequence of genes is proposed: -metJ-metB-metF-argF-thi-purD-metA-metI-metH-. The bearing of these results on the regulation of themetB–FandmetA–Hclusters is discussed.

-2 p~) .Competition studies have shown that the permease has little or no affinity for the other L-amino acids commonly found in proteins. Methionine uptake was competitively inhibited by the growth inhj bitory analogues DLethionine, a-methyl-DL-methionine and DL-methionhe-DL-sulphoximine. Mutants resistant to a-methyl-methionine and methionine sulphoximine have been isolated which were severely defective in the niethionine specific permease. Two of these mutants, metP760 and rnetP761, mapped away from all previously located methionine structural and regulatory genes.

The systems which transport methionine in Salmonella typhimurium LT2 have been studied. Fourteen mutants, isolated by three different selection procedures, had similar growth characteristics and defects in the specific transport process showing a Km of 0 . 3 ,~~ for L-methionine, and therefore lack the high-affinity, metP transport system. The sites of mutation in four of the mutants were shown by P1-mediated transduction to be linked (0-3 to 1.1 %) with a proline marker located at unit 7 on the S. typhimurium chromosome. The high-affinity system was subject to both repression and transinhibition by methionine, and it may also be regulated by the metJ and metK genes. There appeared to be at least two additional transport systems with relatively low affinities for methionine in the metP763 mutant strain, with apparent Km values for methionine of 24 ,UM and approximately 1.8 mM. The latter system, with a very low affinity for methionine, was inhibited by leucine. In addition, methionine inhibited leucine transport, suggesting that one of the low-affinity methionine transport systems may actually be a leucine transport system.

SUMMARYTransport of glutamine by the high-affinity transport system is regulated by the nitrogen status of the medium. With high concentrations of ammonia, transport is repressed; whereas with Casamino acids, transport is elevated, showing behaviour similar to glutamine synthetase . A glutamine auxotroph, lacking glutamine synthetase activity, had elevated transport activity even in the presence of high concentrations of ammonia (and glutamine). This suggests that glutamine synthetase is involved in the regulation of the transport system. A mutant with low glutamate synthase activity had low glutamine transport and glutamine synthetase activities, which could not be derepressed. A mutant in the highaffinity glutamine transport system showed normal regulation of glutamate synthase and glutamine synthetase. Possible mechanisms for this regulation are discussed.