FAD辅助的人源重组静息巯基氧化酶HsQSOX1b活性的测定和功能结构域研究
ctivity detection of Human recombinant quiescent sulfhydryl oxidase HsQSOX1b and study of functional domains
静息巯基氧化酶蛋白(QSOX)是一种黄素依赖型的巯基氧化酶家族蛋白。本文克隆并可溶性表达了无信号肽的人源重组静息巯基氧化酶HsQSOX1b(30-604)及其截短体蛋白。以三(2-羰基乙基)磷盐酸盐(TCEP)、二硫苏糖醇(DTT)和还原性RNA酶(rRNase)为还原底物检测这些蛋白的氧化活力，最终获得了能氧化TCEP的最小结构域蛋白HsQSOX1b(295-556)。由于缺少HsQSOX1b(30-604)全长蛋白的TRX区和C端的部分氨基酸，HsQSOX1b(295-556)蛋白失去了对DTT和rRNase的氧化活力，但保留了对TCEP的氧化活力，HsQSOX1b(295-556)蛋白对TCEP的氧化活力是HsQSOX1b(30-604)全长蛋白氧化活力的42%，且HsQSOX1b(295-556)蛋白比较稳定，可用于蛋白结晶方面的研究。
Quiescent sulfhydryl oxidase(QSOX)is a member of flavin-linked sulfhydryl oxidase family.In this study,HsQSOX1b without signal peptide(HsQSOX1b(30-604)) and its truncated proteins are cloned and soluably expressed,then oxidative activity of these proteins to reduced substrate Tris-(2-carboxyethyl)phosphine hydrochloride(TCEP),dithiothreitol(DTT)and reduced ribonucleases(rRNase) are detected,finally the shortest protein HsQSOX1b(295-556) with oxidative activity to TCEP is confirmed.HsQSOX1b(295-556)is not able to oxidize the reduced small molecule-DTT or the reduced macromolecule rRNase for it lacks TRX domain and part of amino acids of the C-terminal of HsQSOX1b(30-604),while HsQSOX1b(295-556) retains 42% of oxidative activity of HsQSOX1b(30-604)to TCEP,meanwhile,HsQSOX1b(295-556) is stable,so it can be used as a good object of crystallization study.
胡薇、杨弋、张文耀、郑文云
生物化学分子生物学生物工程学
人源重组静息巯基氧化酶截短体蛋白EPrRNase
human quiescin-sulfhydryl oxidasetruncated proteinTCEPDTTrRNase
胡薇,杨弋,张文耀,郑文云.FAD辅助的人源重组静息巯基氧化酶HsQSOX1b活性的测定和功能结构域研究[EB/OL].(2011-05-10)[2025-08-11].http://www.paper.edu.cn/releasepaper/content/201105-234.点此复制
评论