Transactions of the Korean hydrogen and new energy society (한국수소및신에너지학회논문집)

The Korean Hydrogen and New Energy Society (한국수소및신에너지학회)
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Purification of Hydrogenases from Purple Sulfur Bacterium Thiocapsa roseopersicina Using Various Applications of Chromatography

홍색 유황세근 Thiocapsa roseopersicina로 부터 생산되는 Hydrogenase의 각종 크로마토그래피에 의한 정제

  • 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)
  • 최은혜 (한국에너지기술연구원 바이오에너지연구센터) ;
  • 오유관 (한국에너지기술연구원 바이오에너지연구센터) ;
  • 김미선 (한국에너지기술연구원 바이오에너지연구센터)
  • Published : 2008.04.29
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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

References

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