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. 2017 Apr 24;8:15009. doi: 10.1038/ncomms15009

Figure 4. Mutational effects on the transport properties of VcINDY.

Figure 4

(a) The initial rates of succinate transport at 200 mM external Na+ and pH 7.4 were plotted against external succinate concentrations. The data for S146A were averaged to fit to the Michaelis–Menten equation, yielding a KM of 2.57 mM and a Vmax of 416 nmol mg−1 min−1. Transport for T373A remained unsaturable up to 20 mM succinate, suggesting a KM of >270 mM and precluding an accurate measurement of KM or Vmax. (b) The initial rates of succinate transport at 200 μM external substrate concentration (∼8% of the KM for S146A) and pH 7.4 were plotted against the common logarithm of external Na+ concentrations. The data for S146A were averaged to fit to the Hill equation, yielding a Hill coefficient of 3.2 and a K0.5-Na of 36.9 mM. T373A-mediated transport of succinate lacked Na+-dependence within the tested range (up to 800 mM), suggesting that the K0.5-Na for T373A is >800 mM. Similar observations were made with 500 μM (∼20% of the KM) and 4 mM external succinate for S146A and T373A, respectively. At 500 μM succinate, the data for S146A revealed a Hill coefficient of 3.1 and a K0.5-Na of 47.6 mM. For comparison, the data for VcINDY were plotted to highlight the functional consequences of mutations in Na1 and Na2.