Characterization of the denaturation and renaturation of human plasma vitronectin. I. Biophysical characterization of protein unfolding and multimerization

Upon treatment with denaturing agents, vitronectin has been observed to exhibit conformational alterations which are similar to the structural changes detected when vitronectin binds the thrombin-antithrombin complex or associates with the terminal attack complex of complement. Denaturation and rena...

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Bibliographic Details
Published in:The Journal of biological chemistry 1996-06, Vol.271 (24), p.14323-14332
Main Authors: Zhuang, P, Blackburn, M N, Peterson, C B
Format: Article
Language:English
Subjects:
Chromatography, High Pressure Liquid
Circular Dichroism
Guanidine
Guanidines - pharmacology
Humans
Kinetics
Macromolecular Substances
Models, Theoretical
Molecular Weight
Protein Conformation
Protein Denaturation
Protein Folding
Spectrometry, Fluorescence
Ultracentrifugation
Urea - pharmacology
Vitronectin - blood
Vitronectin - chemistry
Vitronectin - isolation & purification
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Summary:Upon treatment with denaturing agents, vitronectin has been observed to exhibit conformational alterations which are similar to the structural changes detected when vitronectin binds the thrombin-antithrombin complex or associates with the terminal attack complex of complement. Denaturation and renaturation of vitronectin isolated from human plasma were characterized by changes in intrinsic fluorescence. Unfolding by chemical denaturants was irreversible and accompanied by self-association of the protein to form vitronectin multimers. Self-association was evaluated by equilibrium analytical ultracentrifugation which demonstrated that multimers form only during the refolding process after removal of denaturant, that multimeric vitronectin dissociates to constituent subunits readily upon treatment with chemical denaturant, and that intermolecular disulfide cross-linking occurs primarily at the dimer level among a subset of constituent vitronectin subunits within the multimer. The monomeric form of vitronectin isolated from human plasma partially unfolds at intermediate concentrations of denaturant to an altered conformation with a high propensity to associate into multimers. Folding of vitronectin in vivo appears to be regulated by partitioning of folding intermediates toward either of two conformations, one that exists as a stable monomer and another that associates into a multimeric form.
ISSN:0021-9258
DOI:10.1074/jbc.271.24.14323