MBA4 was grown in minimal medium containing acetate (squares) or MCA (circles). Uptakes of 50 μM of [2-14C]acetate were assayed in the presence of 0, 5, 10, 25, and 50 μM of CCCP for a period of 1 min. Data shown are the means of three independent experiments, and the error bars represent the standard deviations. A limitation of SGC-CBP30 mouse employing CCCP is that
it cannot discriminate selleck inhibitor between proton-coupled symport and Na+-coupled symport [17, 20]. As it is difficult to remove sodium from the buffers completely and radioactive MCA and acetate were provided in the form of a sodium salt, the effect of pH on acetate- and MCA- uptake was examined with an aim to find out the possible involvement of proton(s). In acetate uptake of acetate-grown cells, the uptake rate decreased steadily as pH increased from 4 to 8 (Figure 5, squares). In acetate uptake of MCA-grown cells, the uptake rate increased slightly as pH increased from 4 to 5 and then dropped gradually as pH increases (Figure 5, circles). The uptake rates were much lower than that of acetate-grown cells in similar assay conditions. In MCA uptake of MCA-grown cells, the uptake rate increased slightly as pH increased from 4 to 6 and dropped swiftly from pH 7 to 8 (Figure 5, triangles). These results showed that acetate- and MCA- transport systems have
different sensitivities to pH. Nonetheless, the involvement of proton(s) in acetate transport is noticeable. Figure 5 Effect of pH on acetate- and MCA- uptake. MBA4 was grown in minimal Saracatinib mouse medium containing acetate or MCA, harvested and resuspended in potassium phosphate buffers of various pH values. Uptakes of 50 μM of [2-14C] labelled acetate or MCA were assayed for a period of 1 min. Squares represent acetate uptake of acetate-grown cells, circles represent acetate uptake of MCA-grown cells, and triangles represent MCA uptake of MCA-grown cells. Data shown are the means of three independent experiments, and the error bars represent the standard deviations. Discussion In this study, we demonstrated Teicoplanin the presence of distinct acetate- and MCA-
transport system in MBA4. This is supported by: (i) the observation that the inducible substrates for acetate- and MCA- uptake activity were different; (ii) the two transport systems have different competing solutes and (iii) a difference in dependency on pH for the two systems. The failure of pyruvate-grown cells to take up acetate suggested that the acetate-transport system in MBA4 was inducible. Both acetate and MCA were able to induce acetate-uptake activity although to a different level. Acetate permease MctC of Corynebacterium glutamicum is also inducible. MctC exhibits a high affinity for acetate and propionate and low affinity for pyruvate. In this case, the expression was higher in pyruvate- than in acetate-grown cells. As a result, both pyruvate- and acetate-grown cells showed comparable acetate-uptake activities .