Plasmalemma Dicarboxylate Transporter of Saccharomyces cerevisiae is Involved in Citrate and Succinate Influx and is Modulated by pH and Cations

摘要

Succinate and citrate transport into yeast (Saccharomyces cerevisiae) cells was studied by measur-ing substrate oxidation rates in the presence and in the absence of effective impermeable oxidation inhibitorsO-palmitoyl-L-malate and 2-undecyl malonate. Linearity of the Dixon plot for 2-undecyl malonate suggeststhat this inhibitor blocked the rate-limiting step upon oxidation of both substrates, which was, most probably,transport of these substrates across the plasma membrane (due to inability of the inhibitor to penetrate into themembrane). This approach allowed fast (within 30-40 min) measurement of kinetic parameters of the trans-porter in individual samples without losing control over limiting conditions. In case of succinate transport, thelimiting rate of succinate oxidation (Vmax) depended on pH and increased monotonously from near-zero atpH 4.5 to the maximum level at pH 7.5. At pH 5.5, succinate and citrate transport was insensitive to the proto-nophore FCCP, being activated by Na~+ions and competitively inhibited by 2-undecyl malonate and K~+ions.Values of K, for 2-undecyl malonate were similar for both substrates. These data suggest that citrate and succi-nate influx is mediated by a common plasma membrane transporter. This is not typical of fungi. At pH 6.5, Tris+,K~+and Na~+had no effect on succinate oxidation. In monosodium media pH increase was accompanied by adecrease of succinate Km due to higher proportion of the dianionic form of the substrate. Atypical substratespecificity and mechanisms of functional activity of the dicarboxylate transporter in plasma membrane ofS. cerevisiae are discussed.