Deciphering the molecular structure of cryptolepain in organic solvents

• Published in: Biochimie Vol. 94; no. 2; pp. 310 - 317
• Main Authors: Prasanna Kumari, N.K., Jagannadham, M.V.
• Format: Journal Article
• Language: English
• Published: France Elsevier B.V 01-02-2012
• Subjects: Circular Dichroism; Cryptolepis - chemistry; Guanidine - chemistry; Hydrogen-Ion Concentration; Methanol - chemistry; Molten globule; Plant Proteins - chemistry; Plant Proteins - isolation & purification; Protein Denaturation; Protein Folding; Protein Stability; Protein Structure, Secondary; Protein Structure, Tertiary; Sequential unfolding; Serine protease cryptolepain; Serine Proteases - chemistry; Serine Proteases - isolation & purification; Solutions; Solvents - chemistry; Spectrometry, Fluorescence; Temperature; Thermodynamics; Serine protease cryptolepain; CD; UV; TFE; Sequential unfolding; GuHC; Circular dichroism; Molten globule; Fu
• ISSN: 0300-9084; 1638-6183
• DOI: 10.1016/j.biochi.2011.07.017

Solvent composition plays a major role in stabilizing/destabilizing the forces that are responsible for the native structure of a protein. Often, the solvent composition drives the protein into non-native conformations. Elucidation of such non-native structures provides valuable information about the molecular structure of the protein, which is unavailable otherwise. Inclusion of methanol (non-fluorinated alcohol) or TFE (fluorinated alcohol) in the solvent composition drove cryptolepain, a serine protease and an all-β-protein, into a non-native structure with an enhanced β-sheet or induction of α-helix. These solvents did not much affect cryptolepain under neutral conditions, even at higher concentrations, but the effects were predominant at lower pH, when the protein molecule is under stress. The organic solvent-induced state is partially unfolded with similar characteristics to the molten globule state seen with protein under a variety of conditions. Chemical- or temperature-induced unfolding of cryptolepain in the presence of organic solvent is distinctly different from that in the absence of organic solvent. Such different unfolding provided evidence of two structural variants in the molecular structure of the protein as well as the differential stabilization/destabilization of such structural variants and their sequential unfolding.
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► Cryptolepain is a serine protease isolated from the latex of Cryptolepis buchanani. ► Organic solvents induce molten globule state in cryptolepain under acidic conditions. ► The unfolding of the enzyme in organic solvent reveals the molecular structure. ► Cryptolepain molecule contains two domains which unfold sequentially.