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Equilibrium unfolding of A. niger RNase: pH dependence of chemical and thermal denaturation.
Eur Biophys J. 2011 Aug; 40(8):923-35.EB

Abstract

Equilibrium unfolding of A. niger RNase with chemical denaturants, for example GuHCl and urea, and thermal unfolding have been studied as a function of pH using fluorescence, far-UV, near-UV, and absorbance spectroscopy. Because of their ability to affect electrostatic interactions, pH and chemical denaturants have a marked effect on the stability, structure, and function of many globular proteins. ANS binding studies have been conducted to enable understanding of the folding mechanism of the protein in the presence of the denaturants. Spectroscopic studies by absorbance, fluorescence, and circular dichroism and use of K2D software revealed that the enzyme has α + β type secondary structure with approximately 29% α-helix, 24% β-sheet, and 47% random coil. Under neutral conditions the enzyme is stable in urea whereas GuHCl-induced equilibrium unfolding was cooperative. A. niger RNase has little ANS binding even under neutral conditions. Multiple intermediates were populated during the pH-induced unfolding of A. niger RNase. Urea and temperature-induced unfolding of A. niger RNase into the molten globule-like state is non-cooperative, in contrast to the cooperativity seen with the native protein, suggesting the presence of two parts/domains, in the molecular structure of A. niger RNase, with different stability that unfolds in steps. Interestingly, the GuHCl-induced unfolding of the A state (molten globule state) of A. niger RNase is unique, because a low concentration of denaturant not only induces structural change but also facilitates transition from one molten globule like state (A(MG1)) into another (I(MG2)).

Authors+Show Affiliations

School of Biochemical Engineering, Institute of Technology, Banaras Hindu University, Varanasi, India. ravi_33102000@yahoo.comNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

21611854

Citation

Kumar, Gundampati Ravi, et al. "Equilibrium Unfolding of A. Niger RNase: pH Dependence of Chemical and Thermal Denaturation." European Biophysics Journal : EBJ, vol. 40, no. 8, 2011, pp. 923-35.
Kumar GR, Sharma A, Kumari M, et al. Equilibrium unfolding of A. niger RNase: pH dependence of chemical and thermal denaturation. Eur Biophys J. 2011;40(8):923-35.
Kumar, G. R., Sharma, A., Kumari, M., Jagannadham, M. V., & Debnath, M. (2011). Equilibrium unfolding of A. niger RNase: pH dependence of chemical and thermal denaturation. European Biophysics Journal : EBJ, 40(8), 923-35. https://doi.org/10.1007/s00249-011-0708-1
Kumar GR, et al. Equilibrium Unfolding of A. Niger RNase: pH Dependence of Chemical and Thermal Denaturation. Eur Biophys J. 2011;40(8):923-35. PubMed PMID: 21611854.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Equilibrium unfolding of A. niger RNase: pH dependence of chemical and thermal denaturation. AU - Kumar,Gundampati Ravi, AU - Sharma,Anurag, AU - Kumari,Moni, AU - Jagannadham,Medicherla V, AU - Debnath,Mira, Y1 - 2011/05/25/ PY - 2010/12/13/received PY - 2011/04/27/accepted PY - 2011/04/18/revised PY - 2011/5/26/entrez PY - 2011/5/26/pubmed PY - 2011/11/10/medline SP - 923 EP - 35 JF - European biophysics journal : EBJ JO - Eur Biophys J VL - 40 IS - 8 N2 - Equilibrium unfolding of A. niger RNase with chemical denaturants, for example GuHCl and urea, and thermal unfolding have been studied as a function of pH using fluorescence, far-UV, near-UV, and absorbance spectroscopy. Because of their ability to affect electrostatic interactions, pH and chemical denaturants have a marked effect on the stability, structure, and function of many globular proteins. ANS binding studies have been conducted to enable understanding of the folding mechanism of the protein in the presence of the denaturants. Spectroscopic studies by absorbance, fluorescence, and circular dichroism and use of K2D software revealed that the enzyme has α + β type secondary structure with approximately 29% α-helix, 24% β-sheet, and 47% random coil. Under neutral conditions the enzyme is stable in urea whereas GuHCl-induced equilibrium unfolding was cooperative. A. niger RNase has little ANS binding even under neutral conditions. Multiple intermediates were populated during the pH-induced unfolding of A. niger RNase. Urea and temperature-induced unfolding of A. niger RNase into the molten globule-like state is non-cooperative, in contrast to the cooperativity seen with the native protein, suggesting the presence of two parts/domains, in the molecular structure of A. niger RNase, with different stability that unfolds in steps. Interestingly, the GuHCl-induced unfolding of the A state (molten globule state) of A. niger RNase is unique, because a low concentration of denaturant not only induces structural change but also facilitates transition from one molten globule like state (A(MG1)) into another (I(MG2)). SN - 1432-1017 UR - https://www.unboundmedicine.com/medline/citation/21611854/Equilibrium_unfolding_of_A__niger_RNase:_pH_dependence_of_chemical_and_thermal_denaturation_ L2 - https://dx.doi.org/10.1007/s00249-011-0708-1 DB - PRIME DP - Unbound Medicine ER -