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Differences in the unfolding of procerain induced by pH, guanidine hydrochloride, urea, and temperature.
Biochemistry. 2003 Oct 28; 42(42):12287-97.B

Abstract

The structural and functional aspects along with equilibrium unfolding of procerain, a cysteine protease from Calotropis procera, were studied in solution. The energetic parameters and conformational stability of procerain in different states were also estimated and interpreted. Procerain belongs to the alpha + beta class of proteins. At pH 2.0, procerain exists in a partially unfolded state with characteristics of a molten globule-like state, and the protein is predominantly a beta-sheet conformation and exhibits strong ANS binding. GuHCl and temperature denaturation of procerain in the molten globule-like state is noncooperative, contrary to the cooperativity seen with the native protein, suggesting the presence of two parts in the molecular structure of procerain, possibly domains, with different stability that unfolds in steps. Moreover, tryptophan quenching studies suggested the exposure of aromatic residues to solvent in this state. At lower pH, procerain unfolds to the acid-unfolded state, and a further decrease in the pH drives the protein to the A state. The presence of 0.5 M salt in the solvent composition directs the transition to the A state while bypassing the acid-unfolded state. GuHCl-induced unfolding of procerain at pH 3.0 seen by various methods is cooperative, but the transitions are noncoincidental. Besides, a strong ANS binding to the protein is observed at low concentrations of GuHCl, indicating the presence of an intermediate in the unfolding pathway. On the other hand, even in the presence of urea (8 M), procerain retains all the activity as well as structural parameters at neutral pH. However, the protein is susceptible to unfolding by urea at lower pH, and the transitions are cooperative and coincidental. Further, the properties of the molten globule-like state and the intermediate state are different, but both states have the same conformational stability. This indicates that these intermediates may be located on parallel folding routes of procerain.

Authors+Show Affiliations

Molecular Biology Unit, Institute of Medical Sciences, Banaras Hindu University, Varanasi 221005, India.No affiliation info available

Pub Type(s)

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

Language

eng

PubMed ID

14567690

Citation

Dubey, Vikash Kumar, and M V. Jagannadham. "Differences in the Unfolding of Procerain Induced By pH, Guanidine Hydrochloride, Urea, and Temperature." Biochemistry, vol. 42, no. 42, 2003, pp. 12287-97.
Dubey VK, Jagannadham MV. Differences in the unfolding of procerain induced by pH, guanidine hydrochloride, urea, and temperature. Biochemistry. 2003;42(42):12287-97.
Dubey, V. K., & Jagannadham, M. V. (2003). Differences in the unfolding of procerain induced by pH, guanidine hydrochloride, urea, and temperature. Biochemistry, 42(42), 12287-97.
Dubey VK, Jagannadham MV. Differences in the Unfolding of Procerain Induced By pH, Guanidine Hydrochloride, Urea, and Temperature. Biochemistry. 2003 Oct 28;42(42):12287-97. PubMed PMID: 14567690.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Differences in the unfolding of procerain induced by pH, guanidine hydrochloride, urea, and temperature. AU - Dubey,Vikash Kumar, AU - Jagannadham,M V, PY - 2003/10/22/pubmed PY - 2003/12/3/medline PY - 2003/10/22/entrez SP - 12287 EP - 97 JF - Biochemistry JO - Biochemistry VL - 42 IS - 42 N2 - The structural and functional aspects along with equilibrium unfolding of procerain, a cysteine protease from Calotropis procera, were studied in solution. The energetic parameters and conformational stability of procerain in different states were also estimated and interpreted. Procerain belongs to the alpha + beta class of proteins. At pH 2.0, procerain exists in a partially unfolded state with characteristics of a molten globule-like state, and the protein is predominantly a beta-sheet conformation and exhibits strong ANS binding. GuHCl and temperature denaturation of procerain in the molten globule-like state is noncooperative, contrary to the cooperativity seen with the native protein, suggesting the presence of two parts in the molecular structure of procerain, possibly domains, with different stability that unfolds in steps. Moreover, tryptophan quenching studies suggested the exposure of aromatic residues to solvent in this state. At lower pH, procerain unfolds to the acid-unfolded state, and a further decrease in the pH drives the protein to the A state. The presence of 0.5 M salt in the solvent composition directs the transition to the A state while bypassing the acid-unfolded state. GuHCl-induced unfolding of procerain at pH 3.0 seen by various methods is cooperative, but the transitions are noncoincidental. Besides, a strong ANS binding to the protein is observed at low concentrations of GuHCl, indicating the presence of an intermediate in the unfolding pathway. On the other hand, even in the presence of urea (8 M), procerain retains all the activity as well as structural parameters at neutral pH. However, the protein is susceptible to unfolding by urea at lower pH, and the transitions are cooperative and coincidental. Further, the properties of the molten globule-like state and the intermediate state are different, but both states have the same conformational stability. This indicates that these intermediates may be located on parallel folding routes of procerain. SN - 0006-2960 UR - https://www.unboundmedicine.com/medline/citation/14567690/Differences_in_the_unfolding_of_procerain_induced_by_pH_guanidine_hydrochloride_urea_and_temperature_ DB - PRIME DP - Unbound Medicine ER -