The Experts below are selected from a list of 249 Experts worldwide ranked by ideXlab platform
Gu-gang Chang - One of the best experts on this subject based on the ideXlab platform.
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reversible unfolding of the severe acute respiratory syndrome coronavirus main protease in Guanidinium Chloride
Biophysical Journal, 2007Co-Authors: Hui-ping Chang, Chi-yuan Chou, Gu-gang ChangAbstract:Chemical denaturant sensitivity of the dimeric main protease from severe acute respiratory syndrome (SARS) coronavirus to Guanidinium Chloride was examined in terms of fluorescence spectroscopy, circular dichroism, analytical ultracentrifuge, and enzyme activity change. The dimeric enzyme dissociated at Guanidinium Chloride concentration of <0.4 M, at which the enzymatic activity loss showed close correlation with the subunit dissociation. Further increase in Guanidinium Chloride induced a reversible biphasic unfolding of the enzyme. The unfolding of the C-terminal domain-truncated enzyme, on the other hand, followed a monophasic unfolding curve. Different mutants of the full-length protease (W31 and W207/W218), with tryptophanyl residue(s) mutated to phenylalanine at the C-terminal or N-terminal domain, respectively, were constructed. Unfolding curves of these mutants were monophasic but corresponded to the first and second phases of the protease, respectively. The unfolding intermediate of the protease thus represented a folded C-terminal domain but an unfolded N-terminal domain, which is enzymatically inactive due to loss of regulatory properties. The various enzyme forms were characterized in terms of hydrophobicity and size-and-shape distributions. We provide direct evidence for the functional role of C-terminal domain in stabilization of the catalytic N-terminal domain of SARS coronavirus main protease.
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Reversible Unfolding of the Severe Acute Respiratory Syndrome Coronavirus Main Protease in Guanidinium Chloride
Biophysical journal, 2006Co-Authors: Hui-ping Chang, Chi-yuan Chou, Gu-gang ChangAbstract:Chemical denaturant sensitivity of the dimeric main protease from severe acute respiratory syndrome (SARS) coronavirus to Guanidinium Chloride was examined in terms of fluorescence spectroscopy, circular dichroism, analytical ultracentrifuge, and enzyme activity change. The dimeric enzyme dissociated at Guanidinium Chloride concentration of
Hui-ping Chang - One of the best experts on this subject based on the ideXlab platform.
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reversible unfolding of the severe acute respiratory syndrome coronavirus main protease in Guanidinium Chloride
Biophysical Journal, 2007Co-Authors: Hui-ping Chang, Chi-yuan Chou, Gu-gang ChangAbstract:Chemical denaturant sensitivity of the dimeric main protease from severe acute respiratory syndrome (SARS) coronavirus to Guanidinium Chloride was examined in terms of fluorescence spectroscopy, circular dichroism, analytical ultracentrifuge, and enzyme activity change. The dimeric enzyme dissociated at Guanidinium Chloride concentration of <0.4 M, at which the enzymatic activity loss showed close correlation with the subunit dissociation. Further increase in Guanidinium Chloride induced a reversible biphasic unfolding of the enzyme. The unfolding of the C-terminal domain-truncated enzyme, on the other hand, followed a monophasic unfolding curve. Different mutants of the full-length protease (W31 and W207/W218), with tryptophanyl residue(s) mutated to phenylalanine at the C-terminal or N-terminal domain, respectively, were constructed. Unfolding curves of these mutants were monophasic but corresponded to the first and second phases of the protease, respectively. The unfolding intermediate of the protease thus represented a folded C-terminal domain but an unfolded N-terminal domain, which is enzymatically inactive due to loss of regulatory properties. The various enzyme forms were characterized in terms of hydrophobicity and size-and-shape distributions. We provide direct evidence for the functional role of C-terminal domain in stabilization of the catalytic N-terminal domain of SARS coronavirus main protease.
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Reversible Unfolding of the Severe Acute Respiratory Syndrome Coronavirus Main Protease in Guanidinium Chloride
Biophysical journal, 2006Co-Authors: Hui-ping Chang, Chi-yuan Chou, Gu-gang ChangAbstract:Chemical denaturant sensitivity of the dimeric main protease from severe acute respiratory syndrome (SARS) coronavirus to Guanidinium Chloride was examined in terms of fluorescence spectroscopy, circular dichroism, analytical ultracentrifuge, and enzyme activity change. The dimeric enzyme dissociated at Guanidinium Chloride concentration of
Chi-yuan Chou - One of the best experts on this subject based on the ideXlab platform.
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reversible unfolding of the severe acute respiratory syndrome coronavirus main protease in Guanidinium Chloride
Biophysical Journal, 2007Co-Authors: Hui-ping Chang, Chi-yuan Chou, Gu-gang ChangAbstract:Chemical denaturant sensitivity of the dimeric main protease from severe acute respiratory syndrome (SARS) coronavirus to Guanidinium Chloride was examined in terms of fluorescence spectroscopy, circular dichroism, analytical ultracentrifuge, and enzyme activity change. The dimeric enzyme dissociated at Guanidinium Chloride concentration of <0.4 M, at which the enzymatic activity loss showed close correlation with the subunit dissociation. Further increase in Guanidinium Chloride induced a reversible biphasic unfolding of the enzyme. The unfolding of the C-terminal domain-truncated enzyme, on the other hand, followed a monophasic unfolding curve. Different mutants of the full-length protease (W31 and W207/W218), with tryptophanyl residue(s) mutated to phenylalanine at the C-terminal or N-terminal domain, respectively, were constructed. Unfolding curves of these mutants were monophasic but corresponded to the first and second phases of the protease, respectively. The unfolding intermediate of the protease thus represented a folded C-terminal domain but an unfolded N-terminal domain, which is enzymatically inactive due to loss of regulatory properties. The various enzyme forms were characterized in terms of hydrophobicity and size-and-shape distributions. We provide direct evidence for the functional role of C-terminal domain in stabilization of the catalytic N-terminal domain of SARS coronavirus main protease.
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Reversible Unfolding of the Severe Acute Respiratory Syndrome Coronavirus Main Protease in Guanidinium Chloride
Biophysical journal, 2006Co-Authors: Hui-ping Chang, Chi-yuan Chou, Gu-gang ChangAbstract:Chemical denaturant sensitivity of the dimeric main protease from severe acute respiratory syndrome (SARS) coronavirus to Guanidinium Chloride was examined in terms of fluorescence spectroscopy, circular dichroism, analytical ultracentrifuge, and enzyme activity change. The dimeric enzyme dissociated at Guanidinium Chloride concentration of
Vinod Bhakuni - One of the best experts on this subject based on the ideXlab platform.
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Guanidinium Chloride- and urea-induced unfolding of FprA, a mycobacterium NADPH-ferredoxin reductase: stabilization of an apo-protein by GdmCl.
The FEBS journal, 2005Co-Authors: Nidhi Shukla, Anant Narayan Bhatt, Alessandro Aliverti, Giuliana Zanetti, Vinod BhakuniAbstract:The Guanidinium Chloride- and urea-induced unfolding of FprA, a mycobacterium NADPH-ferredoxin reductase, was examined in detail using multiple spectroscopic techniques, enzyme activity measurements and size exclusion chromatography. The equilibrium unfolding of FprA by urea is a cooperative process where no stabilization of any partially folded intermediate of protein is observed. In comparison, the unfolding of FprA by Guanidinium Chloride proceeds through intermediates that are stabilized by interaction of protein with Guanidinium Chloride. In the presence of low concentrations of Guanidinium Chloride the protein undergoes compaction of the native conformation; this is due to optimization of charge in the native protein caused by electrostatic shielding by the Guanidinium cation of charges on the polar groups located on the protein side chains. At a Guanidinium Chloride concentration of about 0.8 m, stabilization of apo-protein was observed. The stabilization of apo-FprA by Guanidinium Chloride is probably the result of direct binding of the Gdm+ cation to protein. The results presented here suggest that the difference between the urea- and Guanidinium Chloride-induced unfolding of FprA could be due to electrostatic interactions stabilizating the native conformation of this protein.
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Guanidinium Chloride- and urea-induced unfolding of the dimeric enzyme glucose oxidase.
Biochemistry, 2002Co-Authors: Sohail Akhtar, Atta Ahmad, Vinod BhakuniAbstract:We have carried out a systematic study on the Guanidinium Chloride- and urea-induced unfolding of glucose oxidase from Aspergillus niger, an acidic dimeric enzyme, using various optical spectroscopic techniques, enzymatic activity measurements, glutaraldehyde cross-linking, and differential scanning calorimetry. The urea-induced unfolding of GOD was a two-state process with dissociation and unfolding of the native dimeric enzyme molecule occurring in a single step. On the contrary, the GdmCl-induced unfolding of GOD was a multiphasic process with stabilization of a conformation more compact than the native enzyme at low GdmCl concentrations and dissociation along with unfolding of enzyme at higher concentrations of GdmCl. The GdmCl-stabilized compact dimeric intermediate of GOD showed an enhanced stability against thermal and urea denaturation as compared to the native GOD dimer. Comparative studies on GOD using GdmCl and NaCl demonstrated that binding of the Gdm+ cation to the enzyme results in stabiliza...
Hai-meng Zhou - One of the best experts on this subject based on the ideXlab platform.
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Comparison of inactivation and unfolding of calf intestinal alkaline phosphatase in Guanidinium Chloride solution
Tsinghua Science and Technology, 2002Co-Authors: Ying-xia Zhang, Hongwei Xi, Hai-meng ZhouAbstract:The changes in activity and unfolding of calf intestinal alkaline phosphatase (CIP) during denaturation in Guanidinium Chloride solutions of different concentrations were investigated using ultraviolet difference absorption spectra and fluorescence emission spectra. Unfolding and inactivation rate constants were measured and compared. The inactivation course is much faster than that of unfolding, which suggests that the active site of CIP containing two zinc ions and one magnesium ion is situated in a limited and flexible region of the enzyme molecule, which is more fragile to the denaturant than the protein as a whole.
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Inactivation and conformational changes of yeast invertase during unfolding in urea and Guanidinium Chloride solutions.
Genomics, 1998Co-Authors: Hai Peng Yang, Hai-meng ZhouAbstract:Yeast invertase exists in two different forms. The cytoplasmic enzyme is non-glycosylated, whereas the external invertase contains approximately 50% carbohydrate of the high mannose type. In this paper, the inactivation and the conformational changes of the yeast external invertase are analyzed for unfolding in urea and Guanidinium Chloride. The results show that much lower concentrations of denaturants are required to bring about inactivation than are required to produce significant conformational changes of the yeast external invertase. The results suggest that the active sites of the external invertase containing carbohydrate residues may display more conformational flexibility than the enzyme molecules as a whole.
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comparison of inactivation and conformational changes of aminoacylase during Guanidinium Chloride denaturation
Biochimica et Biophysica Acta, 1995Co-Authors: Hai-meng Zhou, Hongrui Wang, Tong ZhangAbstract:The inactivation and unfolding of aminoacylase (EC 3.5.1.14) during denaturation by different concentrations of Guanidinium Chloride (GuHCl) have been compared. A marked decrease in enzyme activity is already evident at low GuHCl concentrations before significant unfolding of the enzyme molecule, as monitored by fluorescence, ultraviolet difference absorption and CD measurement. The kinetic theory of the substrate reaction during irreversible inhibition of enzyme activity previously described by Tsou has been applied to a study on the kinetics of the course of inactivation of aminoacylase during denaturation by GuHCl. The inactivation rate constants of free enzyme and substrate-enzyme complex were determined by Tsou's method. The inactivation reaction kinetics were found to be a monophasic first-order reaction. The kinetics of the unfolding were a bisphasic process consisting of two first-order reactions. At lower GuHCl concentration ( 1.0 M), the inactivation rate was too fast to be measured by conventional dynamic methods, whereas the unfolding remained as a bisphasic process with the fast reaction accruing very fast and the slow reaction occurring at a measurable rate. The results suggest that active sites of aminoacylase containing Zn2+ ions are situated in a limited region of the enzyme molecule that is more fragile to denaturants than the protein as a whole.
