Anaerobic expression of Escherichia coli succinate dehydrogenase:: Functional replacement of fumarate reductase in the respiratory chain during anaerobic growth
Anaerobic expression of Escherichia coli succinate dehydrogenase:: Functional replacement of fumarate reductase in the respiratory chain during anaerobic growth
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DOI：
10.1128/jb.180.22.5989-5996.1998
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发表时间：
1998-11-01
影响因子：
3.2
通讯作者：
Cecchini, G
中科院分区：
文献类型：
--
作者：
Maklashina, E;Berthold, DK;Cecchini, G
来自大肠杆菌的琥珀酸-泛醌氧化还原酶（SQR）在需氧生长期间最大程度地表达，此时其催化三羧酸循环中琥珀酸氧化成富马酸并还原膜中的泛醌。该酶在结构和功能上与参与大肠杆菌无氧呼吸的富马酸还原酶（甲基喹啉-富马酸氧化还原酶[QFR]）相似。杆菌富马酸还原酶，这是熟练的琥珀酸氧化，是能够在有氧呼吸的条件下，允许的frdABCD操纵子有氧表达时，功能上取代SQR。SQR以前没有被证明能够支持E.因为酶复合物的表达在很大程度上被厌氧条件抑制。为了获得SQR的厌氧表达，构建了利用与sdhCDAB基因融合的frdABCD操纵子的P-FRD启动子来驱动表达的质粒。结果表明，在富马酸作为末端电子受体的厌氧生长条件下，SQR可以支持E.杆菌在厌氧生长条件下，SQR和QFR的扩增水平相似。从厌氧生长的细胞分离的SQR的催化性能进行了测量，发现是相同的有氧产生的酶。SQR的厌氧表达给出了比在测试条件下从需氧生长的细胞的膜中发现的更大的酶复合物的产率。此外，发现SQR的厌氧表达可使膜用于酶复合物掺入的能力饱和。正如已经看到的扩增的QFR复合物，E。调用容纳通过增加膜的量产生的过量SQR。多余的膜被发现在管状结构，可以看到在薄切片电子显微镜照片。
Succinate-ubiquinone oxidoreductase (SQR) from Escherichia coli is expressed maximally during aerobic growth, when it catalyzes the oxidation of succinate to fumarate in the tricarboxylic acid cycle and reduces ubiquinone in the membrane. The enzyme is similar in structure and function to fumarate reductase (menaquinol-fumarate oxidoreductase [QFR]), which participates in anaerobic respiration by E. coli. Fumarate reductase, which is proficient in succinate oxidation, is able to functionally replace SQR in aerobic respiration when conditions are used to allow the expression of the frdABCD operon aerobically. SQR has not previously been shown to be capable of supporting anaerobic growth of E. coli because expression of the enzyme complex is largely repressed by anaerobic conditions. In order to obtain expression of SQR anaerobically, plasmids which utilize the P-FRD promoter of the frdABCD operon fused to the sdhCDAB genes to drive expression were constructed. It was found that, under anaerobic growth conditions where fumarate is utilized as the terminal electron acceptor, SQR would function to support anaerobic growth of E. coli. The levels of amplification of SQR and QFR were similar under anaerobic growth conditions. The catalytic properties of SQR isolated from anaerobically grown cells were measured and found to be identical to those of enzyme produced aerobically. The anaerobic expression of SQR gave a greater yield of enzyme complex than aas found in the membrane from aerobically grown cells under the conditions tested. In addition, it was found that anaerobic expression of SQR could saturate the capacity of the membrane for incorporation of enzyme complex. As has been seen with the amplified QFR complex, E. call accommodates the excess SQR produced by increasing the amount of membrane. The excess membrane was found in tubular structures that could be seen in thin-section electron micrographs.