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Molecules 2014, 19(1), 233-246; doi:10.3390/molecules19010233
Article

Unfolding Studies of the Cysteine Protease Baupain, a Papain-Like Enzyme from Leaves of Bauhinia forficata: Effect of pH, Guanidine Hydrochloride and Temperature

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Received: 8 October 2013; in revised form: 12 December 2013 / Accepted: 13 December 2013 / Published: 24 December 2013
(This article belongs to the Special Issue Enzyme Chemistry)
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Abstract: Baupain belongs to the α+β class of proteins with a secondary structure-content of 44% α-helix, 16% β-sheet and 12% β-turn. The structural transition induced by pH was found to be noncooperative, with no important differences observed in the pH range from 3.0 to 10.5. At pH 2.0 the protein presented substantial non-native structure with strong ANS binding. Guanidine hydrochloride (GdnHCl)-induced unfolding did not change the protein structure significantly until 4.0 M, indicating the high rigidity of the molecule. The unfolding was cooperative, as seen by the sigmoidal transition curves with midpoints at 4.7 ± 0.2 M and 5.0 ± 0.2 M GdnHCl, as measured by CD and fluorescence spectroscopy. A red shift of 7 nm in intrinsic fluorescence was observed with 6.0 M GdnHCl. Temperature-induced unfolding of baupain was incomplete, and at least 35% of the native structure of the protein was retained, even at high temperature (90 °C). Baupain showed characteristics of a molten globule state, due to preferential ANS binding at pH 2.0 in comparison to the native form (pH 7.0) and completely unfolded (6.0 M GdnHCl) state. Combined with information about N-terminal sequence similarity, these results allow us to include baupain in the papain superfamily.
Keywords: baupain; circular dichroism; cysteine protease; papain; protein conformation baupain; circular dichroism; cysteine protease; papain; protein conformation
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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MDPI and ACS Style

Silva-Lucca, R.A.; Andrade, S.S.; Ferreira, R.S.; Sampaio, M.U.; Oliva, M.L.V. Unfolding Studies of the Cysteine Protease Baupain, a Papain-Like Enzyme from Leaves of Bauhinia forficata: Effect of pH, Guanidine Hydrochloride and Temperature. Molecules 2014, 19, 233-246.

AMA Style

Silva-Lucca RA, Andrade SS, Ferreira RS, Sampaio MU, Oliva MLV. Unfolding Studies of the Cysteine Protease Baupain, a Papain-Like Enzyme from Leaves of Bauhinia forficata: Effect of pH, Guanidine Hydrochloride and Temperature. Molecules. 2014; 19(1):233-246.

Chicago/Turabian Style

Silva-Lucca, Rosemeire A.; Andrade, Sheila S.; Ferreira, Rodrigo S.; Sampaio, Misako U.; Oliva, Maria Luiza V. 2014. "Unfolding Studies of the Cysteine Protease Baupain, a Papain-Like Enzyme from Leaves of Bauhinia forficata: Effect of pH, Guanidine Hydrochloride and Temperature." Molecules 19, no. 1: 233-246.


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