Conformational change in thrombospondin induced by removal of bound Ca2+ A spin label approach

FEBS Letters

Slane, Jean M.K.¹  Lai, Ching-San¹  Mosher, Deane F.² 

[1] National Biomedical ESR Center, Department of Radiology, Medical College of Wisconsin, Milwaukee, WI 53226, USA;Departments of Medicine and Physiological Chemistry, University of Wisconsin, Madison, WI 53706, USA

关键词: Thrombospondin;    ESR;    Spin labeling;    Protein conformation;    TSP;    thrombospondin;    PMSF;    phenylmethylsulfonyl fluoride;    Proxyl;    2;    2;    5;    5-tetramethyl-1-pyrrolidinyloxyl;    DTNB;    5;    5′-dithiobis(2-nitrobenzoic acid);    Tempo;    2;    2;    6;    6-tetramethyl-1-piperidinooxyl;    SDS-PAGE;    SDS-polyacrylamide gel electrophoresis;   

DOI  :  10.1016/0014-5793(88)81157-8

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The effect of removal of Ca²⁺ bound to thrombospondin (TSP) on the protein structure in solution has been investigated using ESR spin-label techniques. A maleimide spin label was selectively attached to the free thiol group presumably near the carboxyl-terminal domain in which Ca²⁺-binding sites are situated. The ESR spectra of spin-labeled TSP showed that the bound label undergoes a relatively fast rotational motion with an effective rotational correlation time in the nanosecond time regimes. Removal of bound Ca²⁺ in TSP by dialyzing spin-labeled TSP from a Ca²⁺-containing buffer into an EDTA-containing buffer resulted in an increase in the mobility of the bound label by a factor of 2.3. The data suggest that EDTA chelation of bound Ca²⁺ in TSP induces a conformational change of TSP at least near the site of spin labeling.

【 授权许可】

Unknown