Actinobacillus succinogenes is a natural succinate producer, which is the result of fumarate respiration. Succinate production from anaerobic growth with C 4 -dicarboxylates requires transporters catalysing uptake and efflux of C 4 -dicarboxylates. Transporter Asuc_1999 (DcuE) found in A. succinogenes belongs to the Dcu family and was considered the main transporter for fumarate respiration. However, deletion of dcuE affected l-malate uptake of A. succinogenes rather than fumarate uptake. DcuE complemented anaerobic growth of Escherichia coli on l-malate or fumarate; thus, the transporter was characterized in E. coli heterologously. Time-dependent uptake and competitive inhibition assays demonstrated that l-malate is the most preferred substrate for uptake by DcuE. The V max of DcuE for l-malate was 20.04 μmol/gDW·min with K m of 57 μM. The V max for l-malate was comparable to that for fumarate, whereas the K m for l-malate was 8 times lower than that for fumarate. The catalytic efficiency of DcuE for l-malate was 7.3-fold higher than that for fumarate, showing high efficiency and high affinity for l-malate. Furthermore, DcuE catalysed the reversible exchange of three C 4 -dicarboxylates – l-malate, fumarate and succinate – but the preferred substrate for uptake was l-malate. Under physiological conditions, the C 4 -dicarboxylates were reduced to succinate. Therefore, DcuE is proposed as the l-malate/succinate antiporter in A. succinogenes.
Bibliographical noteFunding Information:
This work was supported by the Basic Science Research Program and Basic Research Laboratory (BRL) through the National Research Foundation of Korea (NRF-2015R1C1A2A01053608) and (NRF-2015R1A4A1041997). We are grateful to Gottfried Unden (Mainz University, Mainz) for supplying the plasmids for the experiments.
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