Purification and characterization of a highly thermostable, oxygen-resistant, respiratory [NiFe]-hydrogenase from a marine, aerobic hydrogen-oxidizing bacterium Hydrogenovibrio marinus
Purification and characterization of a highly thermostable, oxygen-resistant, respiratory [NiFe]-hydrogenase from a marine, aerobic hydrogen-oxidizing bacterium Hydrogenovibrio marinus
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DOI：
10.1016/j.ijhydene.2011.03.049
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发表时间：
2011-06-01
影响因子：
7.2
通讯作者：
Nishihara, Hirofumi
中科院分区：
文献类型：
--
作者：
Yoon, Ki-Seok;Fukuda, Keiichi;Nishihara, Hirofumi
在厌氧条件下，从海洋氢化弧菌（Hydrogenoibrio marinus，HmMBH）中分离纯化了膜结合型[NiFe]-氢化酶。其分子量估计为110 kDa，由66 kDa和37 kDa亚基的异二聚体结构组成。纯化的酶在较宽的温度范围内表现出较高的活性：30 ℃时为185 U/mg，85 ℃时为615 U/mg（最适温度）。H（2）的Km和k（cat）/K（m）值分别为12 μ M和8.58 × 10（7）M（-1）s（-1）。最适反应pH为7.8，但在pH 4.0-7.0时稳定性特别高。结果表明，HmMBH是显着的热稳定性和耐氧性：其半衰期为75小时，在80 ℃下的H（2），和超过72小时，在4 ℃下的空气。空气氧化72 h的HmMBH仅显示出弱的Ni-B EPR信号，表明活性中心不易被氧化的结构特征。版权所有（C）2011，氢能出版有限责任公司。由爱思唯尔有限公司出版。保留所有权利。
The membrane-bound [NiFe]-hydrogenase from Hydrogenooibrio marinus (HmMBH) was purified homogeneously under anaerobic conditions. Its molecular weight was estimated as 110 kDa, consisting of a heterodimeric structure of 66 kDa and 37 kDa subunits. The purified enzyme exhibited high activity in a wide temperature range: 185 U/mg at 30 degrees C and 615 U/mg at 85 degrees C (the optimum temperature). The Km and k(cat)/K(m) values for H(2) were, respectively, 12 mu M and 8.58 x 10(7) M(-1) s(-1). The optimum reaction pH was 7.8, but its stability was particularly high at pH 4.0-7.0. Results show that HmMBH was remarkably thermostable and oxygen-resistant: its half-life was 75 h at 80 degrees C under H(2), and more than 72 h at 4 degrees C under air. The air-oxidized HmMBH for 72 h showed only weak EPR signals of Ni-B, suggesting a structural feature in which the active center is not easily oxidized. Copyright (C) 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.