Rhie Mi Na, Yoon Hyo Eun, Oh Hye Yun, Zedler Sandra, Unden Gottfried, Kim Ok Bin
Department of Life Sciences, Division of EcoCreative, Ewha Womans University, 120-750 Seoul, Korea.
Institute for Microbiology and Wine Research, Johannes Gutenberg University Mainz, Becherweg 15, 55099 Mainz, Germany.
Microbiology (Reading). 2014 Jul;160(Pt 7):1533-1544. doi: 10.1099/mic.0.076786-0. Epub 2014 Apr 17.
Actinobacillus succinogenes, which is known to produce large amounts of succinate during fermentation of hexoses, was able to grow on C4-dicarboxylates such as fumarate under aerobic and anaerobic conditions. Anaerobic growth on fumarate was stimulated by glycerol and the major product was succinate, indicating the involvement of fumarate respiration similar to succinate production from glucose. The aerobic growth on C4-dicarboxylates and the transport proteins involved were studied. Fumarate was oxidized to acetate. The genome of A. succinogenes encodes six proteins with similarity to secondary C4-dicarboxylate transporters, including transporters of the Dcu (C4-dicarboxylate uptake), DcuC (C4-dicarboxylate uptake C), DASS (divalent anion : sodium symporter) and TDT (tellurite resistance dicarboxylate transporter) family. From the cloned genes, Asuc_0304 of the DASS family protein was able to restore aerobic growth on C4-dicarboxylates in a C4-dicarboxylate-transport-negative Escherichia coli strain. The strain regained succinate or fumarate uptake, which was dependent on the electrochemical proton potential and the presence of Na(+). The transport had an optimum pH ~7, indicating transport of the dianionic C4-dicarboxylates. Transport competition experiments suggested substrate specificity for fumarate and succinate. The transport characteristics for C4-dicarboxylate uptake by cells of aerobically grown A. succinogenes were similar to those of Asuc_0304 expressed in E. coli, suggesting that Asuc_0304 has an important role in aerobic fumarate uptake in A. succinogenes. Asuc_0304 has sequence similarity to bacterial Na(+)-dicarboxylate cotransporters and contains the carboxylate-binding signature. Asuc_0304 was named SdcA (sodium-coupled C4-dicarboxylate transporter from A. succinogenes).
琥珀酸放线杆菌在己糖发酵过程中能产生大量琥珀酸，在有氧和无氧条件下均可利用延胡索酸等C4 - 二羧酸生长。甘油可刺激其在延胡索酸上的厌氧生长，主要产物为琥珀酸，这表明存在类似于由葡萄糖产生琥珀酸的延胡索酸呼吸作用。对其在C4 - 二羧酸上的有氧生长及相关转运蛋白进行了研究。延胡索酸被氧化为乙酸。琥珀酸放线杆菌的基因组编码六种与二级C4 - 二羧酸转运蛋白相似的蛋白质，包括Dcu（C4 - 二羧酸摄取）、DcuC（C4 - 二羧酸摄取C）、DASS（二价阴离子：钠同向转运体）和TDT（抗碲酸盐二羧酸转运体）家族的转运蛋白。从克隆基因来看，DASS家族蛋白Asuc_0304能够恢复C4 - 二羧酸转运阴性的大肠杆菌菌株在C4 - 二羧酸上的有氧生长。该菌株恢复了琥珀酸或延胡索酸的摄取，这取决于电化学质子势和Na(+)的存在。转运的最适pH约为7，表明二价阴离子C4 - 二羧酸的转运。转运竞争实验表明对延胡索酸和琥珀酸具有底物特异性。有氧生长的琥珀酸放线杆菌细胞摄取C4 - 二羧酸的转运特性与在大肠杆菌中表达的Asuc_0304相似，这表明Asuc_0304在琥珀酸放线杆菌有氧摄取延胡索酸过程中起重要作用。Asuc_0304与细菌Na(+) - 二羧酸共转运蛋白具有序列相似性，并含有羧酸盐结合基序。Asuc_0304被命名为SdcA（来自琥珀酸放线杆菌的钠偶联C4 - 二羧酸转运体）。
