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Probing the conformational state of a truncated staphylococcal nuclease R using time of flight mass spectrometry with limited proteolysis
Yang, F; Cheng, Y; Peng, JR; Zhou, JM; Jing, GZ
关键词conformation ligand binding mass spectrometry truncated staphylococcal nuclease
刊名EUROPEAN JOURNAL OF BIOCHEMISTRY
2001-08-01
268期:15页:4227-4232
收录类别SCI
文章类型Article
WOS标题词Science & Technology
类目[WOS]Biochemistry & Molecular Biology
研究领域[WOS]Biochemistry & Molecular Biology
关键词[WOS]N-TERMINAL FRAGMENTS ; RESIDUAL STRUCTURE ; CRYSTAL-STRUCTURE ; MOLTEN GLOBULE ; PROTEINS ; PEPTIDE ; FOLD
英文摘要

The conformational state of C-terminally truncated staphylococcal nuclease R (SNR135), with and without bound ligands, has been studied by performing limited proteolysis with a specific endoproteinase Glu-C followed by electrophoresis and mass spectrometry. Comparison of the accessibility of the cleavage sites shows that the C-terminal truncation of 14 amino-acid residues causes significant unfolding of the C-terminal part of alpha helix 1 and the center of alpha helix 2, but there is little effect on other regions of the nuclease, in particular the N-terminal subdomain, which includes the active site of the nuclease. The truncation also makes the overall conformation of the nuclease more loose and flexible. Binding of ligands makes helices 1 and 2 more resistant to protease Glu-C attack and converts the partially unfolded state to a native-like state, although the conformational stability of the SNR135 complex is still much lower than that of the full-length enzyme. The results suggest that the amino-acid residues around the active site in the truncated nuclease are arranged in a similar topology to those in the full-length nuclease. The study shows that there is a clear-cut correlation between protease susceptibility and conformational stability of the protein, and the initial proteolytic events are the most critical for evaluating the conformational features of the protein. This study demonstrates how mass spectrometry can be combined with limited proteolysis to observe conformational changes induced by ligand binding.

语种英语
WOS记录号WOS:000170399300014
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被引频次:11[WOS]   [WOS记录]     [WOS相关记录]
文献类型期刊论文
条目标识符http://ir.bjmu.edu.cn/handle/400002259/123622
专题北京大学医药卫生分析中心
作者单位1.Acad Sinica, Inst Biophys, Natl Lab Biomacromol, Beijing 100101, Peoples R China
2.Beijing Univ, Med & Hlth Anal Ctr, Beijing 100871, Peoples R China
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Yang, F,Cheng, Y,Peng, JR,et al. Probing the conformational state of a truncated staphylococcal nuclease R using time of flight mass spectrometry with limited proteolysis[J]. EUROPEAN JOURNAL OF BIOCHEMISTRY,2001,268(15):4227-4232.
APA Yang, F,Cheng, Y,Peng, JR,Zhou, JM,&Jing, GZ.(2001).Probing the conformational state of a truncated staphylococcal nuclease R using time of flight mass spectrometry with limited proteolysis.EUROPEAN JOURNAL OF BIOCHEMISTRY,268(15),4227-4232.
MLA Yang, F,et al."Probing the conformational state of a truncated staphylococcal nuclease R using time of flight mass spectrometry with limited proteolysis".EUROPEAN JOURNAL OF BIOCHEMISTRY 268.15(2001):4227-4232.
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