초고열성 고세균 Pyrococcus horikoshii 유래 샤페로닌의 ATPase 활성 특성
Characterization of ATPase Activity of Chaperonin from the Hyperthermophilic Archaeon Pyrococcus horikoshii
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초록
Group II형 샤페로닌은 단백질의 캡슐화를 유도하기 위해열린 기질 결합 형태에서 닫힌 형태로 형태를 변화시키며, 이 때 ATP를 필요로 한다. 샤페로닌의 폴딩 유도는 ATP에의한 샤페로닌의 구조 변화와 관련이 있는 것으로 보여진다. 본 연구에서는 Pyrococcus horikoshii OT3의 group II형샤페로닌인 PhCpn의 ATPase 활성을 다양한 조건에서 측정하였다. PhCpn의 반응온도(37−85℃)와 ATP 농도(1.5− 10 mM) 의존성을 확인한 결과, 반응 온도는 80℃에서, ATP 농도는 3 mM에서 최적 활성을 보였다. 염의 종류에 따른ATPase의 활성을 분석한 결과, 1가 양이온은 300 mM LiCl, 2가 양이온은 5 mM MgCl2에서 최적 활성을 나타내었다. ATP 기질에 대한 Km 값은 2.17mM, Vmax 값은 833.3 μM/ min으로 계산되었다. 이러한 결과는 의약학용 및 바이오 산업용 단백질(효소)을 장기간 활성유지하는데 PhCpn을 이용할 경우에 귀중한 기초 자료를 제공할 것이다.
Abstract
ATP drives the conformational change of the group II chaperonin from the open lid substrate-binding conformation to the closed lid conformation to encapsulate an unfolded protein in the central cavity. It is thought that the folding activity of group II chaperonin is strongly correlated with the ATP-dependent conformational change ability. In order to confirm the dependence of the reaction temperature and ATP concentration of PhCpn, the ATPase activities were measured under different reaction temperatures and ATP concentrations. The maximal ATPase activity of PhCpn was observed at 80℃ and 3 mM ATP concentration. As a result of ATPase activity according to the type of salt ions, the highest activity was observed at 300 mM LiCl among the univalent cations and 5 mM MgCl2 among the divalent cations, respectively. The values of Km and Vmax for ATP substrate were estimated as 2.17 mM and 833.3 μM/min, respectively. This results provide the enzymatic information of PhCpn when the prolonged and high activities of pharmaceutical and industrial proteins (or enzymes), by using chaperonin molecules, are required.
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