CAJ | 학술논문

为了对人源含硒单链抗体酶Se-scFv-B3的底物结合部位和催化基团进行研究,在理论预测的基础上,通过快速定点突变法分别在2个理论预测的底物结合部位(位点1和位点2)内选定Ala180和Ala44定点突变为丝氨酸(Ser).2个突变体蛋白经化学修饰将Ser转变成谷胱甘肽过氧化物酶(GPX)的催化基团硒代半胱氨酸(Sec)后,前者的GPX活力达到了Se-scFv-B3的2倍多,而后者的GPX活力没有明显提高,这表明位点1可能是主要的底物结合部位,与理论预测的结果一致.
위료대인원함서단련항체매Se-scFv-B3적저물결합부위화최화기단진행연구,재이론예측적기출상,통과쾌속정점돌변법분별재2개이론예측적저물결합부위(위점1화위점2)내선정Ala180화Ala44정점돌변위사안산(Ser).2개돌변체단백경화학수식장Ser전변성곡광감태과양화물매(GPX)적최화기단서대반광안산(Sec)후,전자적GPX활력체도료Se-scFv-B3적2배다,이후자적GPX활력몰유명현제고,저표명위점1가능시주요적저물결합부위,여이론예측적결과일치.
Ala180 in site 1 and Ala44 in site 2 of human selenium-containing single chain abzyme Se-scFv-B3 were chosen to be mutated to serines, respectively, to study the substrate binding sites and the catalytic group of Se-scFv-B3 and to find the reason why the Se-scFv-B3 has lower catalytic activity based on the theoretical anticipation for the protein. After serine residue was reacted into secystins by chemical modification, the activity of the anterior one is about two times than that of the Se-scFv-B3, but the latter one did not offer a significant improvement. This indicated that site 1 was the predominant binding site for GSH, which was consistent with the foregoing conjecture and the results of energy calculation. In this study, the GPX activity of Se-scFv-B3 was improved and the site 1 preliminarily confirmed to be the substrate binding site based on both theoretical and experimental research. All of these observations will be useful for further work in this area.