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The Scientific World Journal
Volume 2014, Article ID 824768, 9 pages
http://dx.doi.org/10.1155/2014/824768
Research Article

Molten Globule-Like Partially Folded State of Bacillus licheniformis α-Amylase at Low pH Induced by 1,1,1,3,3,3-Hexafluoroisopropanol

1Biomolecular Research Group, Biochemistry Programme, Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia
2Faculty of Applied Medical Sciences, University of Gezira, P.O. Box 20, 2667 Wad Medani, Sudan

Received 3 January 2014; Accepted 17 February 2014; Published 7 April 2014

Academic Editors: B. Sharma and A. Surguchov

Copyright © 2014 Adyani Azizah Abd Halim et al. 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.

Abstract

Effect of 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) on acid-denatured Bacillus licheniformis α-amylase (BLA) at pH 2.0 was investigated by far-UV CD, intrinsic fluorescence, and ANS fluorescence measurements. Addition of increasing HFIP concentrations led to an increase in the mean residue ellipticity at 222 nm () up to 1.5 M HFIP concentration beyond which it sloped off. A small increase in the intrinsic fluorescence and a marked increase in the ANS fluorescence were also observed up to 0.4 M HFIP concentration, both of which decreased thereafter. Far- and near-UV CD spectra of the HFIP-induced state observed at 0.4 M HFIP showed significant retention of the secondary structures closer to native BLA but a disordered tertiary structure. Increase in the ANS fluorescence intensity was also observed with the HFIP-induced state, suggesting exposure of the hydrophobic clusters to the solvent. Furthermore, thermal denaturation of HFIP-induced state showed a non-cooperative transition. Taken together, all these results suggested that HFIP-induced state of BLA represented a molten globule-like state at pH 2.0.