A PRIMARY RESPIRATORY NA-+ PUMP OF AN ANAEROBIC BACTERIUM - THE NA-+-DEPENDENT NADH - QUINONE OXIDOREDUCTASE OF KLEBSIELLA PNEUMONIAE
A PRIMARY RESPIRATORY NA-+ PUMP OF AN ANAEROBIC BACTERIUM - THE NA-+-DEPENDENT NADH - QUINONE OXIDOREDUCTASE OF KLEBSIELLA PNEUMONIAE
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DOI：
10.1007/bf00416604
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发表时间：
1989-04-01
影响因子：
2.8
通讯作者：
THOMER, A
中科院分区：
文献类型：
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作者：
DIMROTH, P;THOMER, A
在柠檬酸盐厌氧条件下生长的肺炎克雷伯菌膜含有 NADH 氧化酶活性，该活性可被 Na+ 或 Li+ 离子特异性激活，并被 2-庚基-4-羟基喹啉-N-氧化物 (HQNO) 有效抑制。细胞色素 b 和 d 存在于膜中，并且细胞色素 b 的稳态还原水平随着 NaCl 的添加而增加。反向细菌膜囊泡在 NADH 氧化后积累 Na+ 离子。 Na + 的吸收被莫能菌素和 HQNO 完全抑制，并被羰基氰化物-对三氟甲氧基苯腙 (FCCP) 轻微刺激，从而表明主 Na + 泵的运行。细菌膜的 Triton 提取物不通过 O2 催化 NADH 氧化，而是通过铁氰化物或甲萘醌以不依赖 Na+ 的方式催化 NADH 氧化。通过添加微摩尔浓度的泛醌-1 可以恢复 O2 对 Na+ 依赖的 NADH 氧化。用 KCN 抑制终止氧化酶后，泛醌-1 和 NADH 形成泛醇。该反应被 10 mM NaCl 刺激约 6 倍，并被少量 HQNO 严重抑制。电子从 NADH 转移到泛醌-1 期间形成超氧自由基。添加 NaCl 后这些自由基会消失，但 NaCl 和 HQNO 不会消失。据认为，超氧自由基是由半醌自由基产生的，半醌自由基是由醌自由基的一个电子还原形成的，而醌自由基是由醌的一个电子还原在Na+独立反应序列中形成的，然后在Na+和HQNO敏感反应中歧化为醌和醌醇。肺炎克雷伯菌呼吸Na+泵的机制似乎与溶藻弧菌非常相似。
Membranes of Klebsiella pneumoniae, grown anaerobically on citrate, contain a NADH oxidase activity that is activated specifically by Na+ or Li+ ions and effectively inhibited by 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO). Cytochromes b and d were present in the membranes, and the steady state reduction level of cytochrome b increased on NaCl addition. Inverted bacterial membrane vesicles accumulated Na+ ions upon NADH oxidation. Na+ uptake was completely inhibited by monensin and by HQNO and slightly stimulated by carbonylcyanide-p-trifluoromethoxy phenylhydrazone (FCCP), thus indicating the operation of a primary Na+ pump. A Triton extract of the bacterial membranes did not catalyze NADH oxidation by O2, but by ferricyanide or menadione in a Na+-independent manner. The Na+-dependent NADH oxidation by O2 was restored by adding ubiquinone-1 in micromolar concentrations. After inhibition of the terminant oxidase with KCN, ubiquinol was formed from ubiquinone-1 and NADH. The reaction was stimulated about 6-fold by 10 mM NaCl and was severely inhibited by low amounts of HQNO. Superoxide radicals were formed during electron transfer from NADH to ubiquinone-1. These radicals disappeared by adding NaCl, but not with NaCl and HQNO. It is suggested that the superoxide radicals arise from semiquinone radicals which are formed by one electron reduction of quinone radicals which are formed by one electron reduction of quinone in a Na+-independent reaction sequence and then dismutate in a Na+ and HQNO sensitive reaction to quinone and quinol. The mechanism of the respiratory Na+ pump of K. pneumoniae appears to be quite similar to that of Vibrio alginolyticus.