| In vitro renaturation of alkaline family G/11 xylanase via a folding intermediate: alpha-crystallin facilitates refolding in an ATP-independent manner. | |
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MedLine Citation:
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PMID: 20703955 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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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. |
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Authors:
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Tanmay Dutta; Arindam Bhattacharjee; Uddalak Majumdar; Saugata Sinha Ray; Rupam Sahoo; Sanjay Ghosh |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2009-11-22 |
Journal Detail:
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Title: Applied biochemistry and biotechnology Volume: 162 ISSN: 1559-0291 ISO Abbreviation: Appl. Biochem. Biotechnol. Publication Date: 2010 Nov |
Date Detail:
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Created Date: 2010-08-12 Completed Date: 2010-11-22 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 8208561 Medline TA: Appl Biochem Biotechnol Country: United States |
Other Details:
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Languages: eng Pagination: 1238-48 Citation Subset: IM |
Affiliation:
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Department of Biochemistry, University College of Science, Calcutta University, Kolkata 700 019, West Bengal, India. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Adenosine Triphosphate
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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* |
| Chemical | |
Reg. No./Substance:
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0/Alkalies; 0/alpha-Crystallins; 113-00-8/Guanidine; 56-65-5/Adenosine Triphosphate; EC 3.2.1.8/Endo-1,4-beta Xylanases |
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