Influence of fluoro, chloro and alkyl alcohols on the folding pathway of human serum albumin.
Urea-induced equilibrium unfolding of human serum albumin (HSA) when studied by mean residue ellipticity at 222 nm (MRE(222)) or intrinsic fluorescence measurements showed a two-step, three-state transition with a stable intermediate around 4.6-5.2 M urea. The presence of 2,2,2-trifluoroethanol (TFE) resulted in a single-step, two-state transition with a significant shift towards higher urea concentration, suggesting the stabilizing effect of TFE. The free energy of stabilization (DeltaDeltaG(D)(H(2)O)) in the presence of 3.0 M TFE was determined to be 2.68 and 2.72 kcal/mol by MRE(222) and fluorescence measurements, respectively. The stabilizing potential of other alcohols on the refolding behavior of HSA at 5.0 M urea (where the intermediate exists) as studied by MRE(222) and intrinsic fluorescence measurements showed the following order: 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) > TFE > 2-chloroethanol > tert-butanol > iso-propanol > ethanol > methanol. Further, the extent of refolding at the highest concentration of alcohol was similar in all cases. The stabilizing effect of TFE on guanidine hydrochloride (GdnHCl)-induced unfolding of HSA was nearly equal to that found for urea denaturation, as reflected in the DeltaDeltaG(D)(H(2)O) value (2.38 kcal/mol). Taken together, these results suggest that the stabilizing effect of TFE and other alcohols on urea/GdnHCl-induced unfolding of HSA is higher for alcohols that contain bulky groups or fluorine atoms.
Kumar
Yogesh
Y
Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh 202 002, India.
Muzammil
Salman
S
Tayyab
Saad
S
D000438:Alcohols / Q000737:chemistry / Q000494:pharmacology*; D002942:Circular Dichroism; D006801:Humans; D011487:Protein Conformation; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D020673:Protein Renaturation / Q000187:drug effects*; D012709:Serum Albumin / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014508:Urea / Q000494:pharmacology
D000438:Alcohols; D012709:Serum Albumin; D014508:Urea