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KMID : 0545120030130050717
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Journal of Microbiology and Biotechnology
2003 Volume.13 No. 5 p.717 ~ p.724
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Oxygen Sensitivity of Carbon Monoxide-Dependent Hydrogen Production Activity in Citrobacter sp.
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KIM, JUNG RAE
OH, YOU-KWAN/YOON, YEO-JOON/LEE, EUN YEOL/PARK, SUNGHOON
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Abstract
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A newly isolated Citrobacter sp. Y19 catalyzes the CO-dependent H, production (biological water-gas shift reaction) by the actions of CO dehydrogenase (CODH) and hydrogenase. Y19 requires 0, for fast growth, but its H₂ production activity is significantly inhibited by 0,. In the present study, the effect of O₂ on the activities of CODH and hydrogenase was investigated quantitatively in both whole cells and broken cells, based on CO-dependent or methyl viologen (MV)-dependent H₂, production in addition to CO-dependent MV reduction. In crude cell extracts, CODH activity was mostly found in the soluble fraction. Inactivation of CODH and hydrogenase activities by 0, followed the first-order decay kinetics, and the dependence of the rate constants on 0, partial pressure could be expressed by the Michaelis-Menten equation. In whole cells, the maximum deactivation rate constants (k_(d max)) of hydrogenase and CODH were quite similar: 0.07±0.03 min¹ and 0.10±0.04 min¹, respectively. However, the first-order rate constant (k,_(max)/K_(?)) of CODH (0.25 min^(-1) atm¹) at low 0₂ partial pressures was about 3-fold higher than that of the hydrogenase, since the half-saturation constant (K,) of CODH was about half of that of hydrogenase. In broken cells, both enzymes became significantly more sensitive to O₂ compared to the unbroken cells, while k_(d max)/K, increased 37-fold for hydrogenase and 6.7-fold for CODH. When whole cells were incubated under anaerobic conditions after being exposed to air for 1 h, hydrogenase activity was recovered more than 90% in 2 h suggesting that the deactivation of hydrogenase by O₂ was reversible. On the contrary, CODH activity was not recovered once deactivated by O₂, and the only way to recover the activity was to synthesize new CODH. This study indicates that O₂ sensitivity of H₂ production activity of Citrobacter sp. Y19 is an important drawback as in other H₂-producing bacteria.
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