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Calcium-ion-induced stabilization of the protease from Bacillus cereus WQ9-2 in aqueous hydrophilic solvents: effect of calcium ion binding on the hydration shell and intramolecular interactions.
MedLine Citation:
PMID:  23322168     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
The neutral protease WQ from Bacillus cereus is stable in various aqueous organic mixtures, with the exception of those containing acetonitrile (ACN) and dimethylformamide (DMF). The stability of the enzyme in aqueous hydrophilic solvents was dramatically enhanced with the addition of calcium ions, with the degree of improvement in the half-life relative to different solutions ranging from fourfold to more than 70-fold. Studies of the kinetic constants showed that calcium ions induced slight conformational changes in the active site of the enzyme in aqueous ACN. We investigated the molecular mechanisms underlying this stabilizing effect by employing a combination of biophysical techniques and molecular dynamics simulation. In aqueous ACN, the intrinsic fluorescence and circular dichroism analysis demonstrated that the addition of calcium ions induced a relatively compact conformation and maintained both the native-like microenvironment near the tryptophan residues and the secondary structure. Alternatively, homology modeling confirmed the location of four calcium-ion-binding sites in the enzyme, and molecular dynamics simulation revealed that three other calcium ions were bound to the surface of the enzyme. Calcium ions, known as a type of kosmotrope, can strongly bond with water molecules, thus aiding in the formation of the regional hydration shell required for the maintenance of enzyme activity. In addition, the introduction of calcium ions resulted in the formation of additional ionic interactions, providing propitious means for protein stabilization. Thus, the stronger intramolecular interactions were also expected to contribute partially to the enhanced stability of the enzyme in an aqueous organic solvent.
Authors:
Jiaxing Xu; Yu Zhuang; Bin Wu; Long Su; Bingfang He
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2013-1-16
Journal Detail:
Title:  Journal of biological inorganic chemistry : JBIC : a publication of the Society of Biological Inorganic Chemistry     Volume:  -     ISSN:  1432-1327     ISO Abbreviation:  J. Biol. Inorg. Chem.     Publication Date:  2013 Jan 
Date Detail:
Created Date:  2013-1-16     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9616326     Medline TA:  J Biol Inorg Chem     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Affiliation:
College of Biotechnology and Pharmaceutical Engineering, Nanjing University of Technology, 30 Puzhunan Road, Nanjing, 211816, Jiangsu, China.
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