Purification and characterization of succinate:menaquinone oxidoreductase from Corynebacterium glutamicum.
Succinate:menaquinone oxidoreductase from Corynebacterium glutamicum, a high-G+C, Gram-positive bacterium, was purified to homogeneity. The enzyme contained two heme B molecules and three polypeptides with apparent molecular masses of 67, 29 and 23 kDa, which corresponded to SdhA (flavoprotein), SdhB (iron-sulfur protein), and SdhC (membrane anchor protein), respectively. In non-denaturating polyacrylamide gel electrophoresis, the enzyme migrated as a single band with an apparent molecular mass of 410 kDa, suggesting that it existed as a trimer. The succinate dehydrogenase activity assayed using 2,3-dimethoxy-5-methyl-6-decyl-1,4-benzoquinone and 2,6-dichloroindophenol as the electron acceptor was inhibited by 2-n-heptyl-4-hydroxyquinoline N-oxide (HQNO), and the Dixon plots were biphasic. In contrast, the succinate dehydrogenase activity assayed using phenazine methosulfate and 2,6-dichloroindophenol was inhibited by p-benzoquinone and not by HQNO. These findings suggested that the C. glutamicum succinate:menaquinone oxidoreductase had two quinone binding sites. In the phylogenetic tree of SdhA, Corynebacterium species do not belong to the high-G+C group, which includes Mycobacterium tuberculosis and Streptomyces coelicolor, but are rather close to the group of low-G+C, Gram-positive bacteria such as Bacillus subtilis. This situation may have arisen due to the horizontal gene transfer.
Kurokawa
Tatsuki
T
Department of Bioscience and Bioinformatics, Kyushu Institute of Technology, Kawazu 680-4, Iizuka, Fukuoka-ken, 820-8502, Japan.
Sakamoto
Junshi
J
D016227:Benzoquinones / Q000494:pharmacology; D048230:Corynebacterium glutamicum / Q000201:enzymology*; D042963:Electron Transport Complex II / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D006418:Heme / Q000032:analysis; D006912:Hydroxyquinolines / Q000494:pharmacology; D010802:Phylogeny
D016227:Benzoquinones; D006912:Hydroxyquinolines; C049991:MY 12-62c; C004532:quinone; D006418:Heme; D042963:Electron Transport Complex II