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Purification of Hydrogenases from Purple Sulfur Bacterium Thiocapsa roseopersicina Using Various Applications of Chromatography  

Choi, Eun-Hye (Bioenergy Research Center, Korea Institute of Energy Research)
Oh, You-Kwan (Bioenergy Research Center, Korea Institute of Energy Research)
Kim, Mi-Sun (Bioenergy Research Center, Korea Institute of Energy Research)
Publication Information
Transactions of the Korean hydrogen and new energy society / v.19, no.2, 2008 , pp. 124-131 More about this Journal
Abstract
Crude cytoplasmic fraction of phototrophic purple sulfur bacterium, Thiocapsa roseopersicina NCIB 8347, were initially prepared and purified by sonication, ultracentrifugation, ammonium sulfate fractionation and heat-treatment and it has been previously reported. Using various applications of chromatography far the purification of membrane-bound and soluble hydrogenases from heat-treated enzyme fraction were studied at present report. When the heat-treated enzyme preparation was applied to the anion column chromatography using Q-sepharose, Fraction I and II, which were extracted with the KCl 0-0.5 M gradient, showed the specific evolution hydrogenase activity 3.86 and 2.27 U/mg-protein respectively. Specific hydrogenase activitys of Fraction I and II were further increased to 4.35 and 7.46 U/mg-protein for Fraction I and to 2.49 and 4.41 U/mg-protein fur Fraction II respectively, when hydrophobic interaction column, Phenyl superose, and anion exchange column, Mono-Q, were applied. Size exclusion chromatography using superdex 200 concentrated the hydrogenase Fraction I and II to 9.19 and 7.84 U/mg-protein respectively at the final step of purification.
Keywords
thiocapsa roseopersicina; hydrogenase; specific hydrogenase activity;
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Times Cited By KSCI : 1  (Citation Analysis)
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