期刊
BIOCHEMICAL JOURNAL
卷 374, 期 -, 页码 657-665出版社
PORTLAND PRESS
DOI: 10.1042/BJ20030642
关键词
formaldehyde dehydrogenase; mycobacteria; mycothiol; S-nitrosomycothiol reductase
When grown in culture Mycobacterium smegmatis metabolized S-nitrosoglutathione to oxidized glutathione and nitrate, which suggested a possible involvement of an S-nitrosothiol reductase and mycobacterial haemoglobin. The mycothiol-dependent formaldehyde dehydrogenase from M. smegmatis was purified by a combination of Ni2+-IMAC (immobilized metal ion affinity chromatography), hydrophobic interaction, anion-exchange and affinity chromatography. The enzyme had a subunit molecular mass of 38 263 kDa. Steady-state kinetic studies indicated that the enzyme catalyses the NAD(+)-dependent conversion of S-hydroxymethylmycothiol into formic acid and mycothiol by a rapid-equilibrium ordered mechanism. The enzyme also catalysed an NADH-dependent decomposition of S-nitrosomycothiol (MSNO) by a sequential mechanism and with an equimolar stoichiometry of NADH:MSNO, which indicated that the enzyme reduces the nitroso group to the oxidation level of nitroxyl. V-max for the MSNO reductase reaction indicated a turnover per subunit of approx. 116 700 min(-1), which was 76-fold faster than the formaldehyde dehydrogenase activity. A gene, Rv2259. annotated as a class III alcohol dehydrogenase in the Mycobacterium tuberculosis genome was cloned and expressed in M. smegmatis as the C-terminally His(6)-tagged product. The purified recombinant enzyme from M. tuberculosis also catalysed both activities. M. smegmatis S-nitrosomycothiol reductase converted MSNO into the N-hydroxysulphenamide, which readily rearranged to mycothiolsulphinamide. In the presence of MSNO reductase, M. tuberculosis HbN (haemoglobin N) was converted with low efficiency into metHbN [HbN(Fe3+)] and this conversion was dependent on turnover of MSNO reductase. These observations suggest a possible route in vivo for the dissimilation of S-nitrosoglutathione.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据