Document Detail

In vitro renaturation of alkaline family G/11 xylanase via a folding intermediate: alpha-crystallin facilitates refolding in an ATP-independent manner.
MedLine Citation:
PMID:  20703955     Owner:  NLM     Status:  MEDLINE    
In this study, alkaliphilic family G/11 xylanase from alkali-tolerant filamentous fungi Penicillium citrinum MTCC 6489 was used as a model system to gain insight into the molecular aspects of unfolding/refolding of alkaliphilic glycosyl hydrolase protein family. The intrinsic protein fluorescence suggested a putative intermediate state of protein in presence of 2 M guanidium hydrochloride (GdmCl) with an emission maximum of 353 nm. Here we studied the refolding of GdmCl-denatured alkaline xylanase in the presence and the absence of a multimeric chaperone protein alpha-crystallin to elucidate the molecular mechanism of intramolecular interactions of the alkaliphilic xylanase protein that dictates its extremophilic character. Our results, based on intrinsic tryptophan fluorescence and hydrophobic fluorophore 8-anilino-1- naphthalene sulfonate-binding studies, suggest that alpha-crystallin formed a complex with a putative molten globule-like intermediate in the refolding pathway of xylanase in an ATP-independent manner. A 2 M GdmCl is sufficient to denature alkaline xylanase completely. The hydrodynamic radius (R(H)) of a native alkaline xylanase is 4.0, which becomes 5.0 in the presence of 2 M GdmCl whereas in presence of the higher concentration of GdmCl R(H) value was shifted to 100, indicating the aggregation of denatured xylanase. The alpha-crystallin.xylanase complex exhibited the recovery of functional activity with the extent of approximately 43%. Addition of ATP to the complex did not show any significant effect on activity recovery of the denatured protein.
Tanmay Dutta; Arindam Bhattacharjee; Uddalak Majumdar; Saugata Sinha Ray; Rupam Sahoo; Sanjay Ghosh
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2009-11-22
Journal Detail:
Title:  Applied biochemistry and biotechnology     Volume:  162     ISSN:  1559-0291     ISO Abbreviation:  Appl. Biochem. Biotechnol.     Publication Date:  2010 Nov 
Date Detail:
Created Date:  2010-08-12     Completed Date:  2010-11-22     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  8208561     Medline TA:  Appl Biochem Biotechnol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1238-48     Citation Subset:  IM    
Department of Biochemistry, University College of Science, Calcutta University, Kolkata 700 019, West Bengal, India.
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MeSH Terms
Adenosine Triphosphate / pharmacology
Alkalies / metabolism*
Endo-1,4-beta Xylanases / metabolism*
Enzyme Activation / drug effects
Guanidine / pharmacology
Hydrophobic and Hydrophilic Interactions
Protein Conformation / drug effects
Protein Denaturation / drug effects
Protein Folding / drug effects*
Protein Renaturation / drug effects*
Spectrometry, Fluorescence
alpha-Crystallins / pharmacology*
Reg. No./Substance:
0/Alkalies; 0/alpha-Crystallins; 113-00-8/Guanidine; 56-65-5/Adenosine Triphosphate; EC,4-beta Xylanases

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