Structural perturbation of proteins in low denaturant concentrations.
The presence of very low concentrations of the widely used chemical denaturants, guanidinium chloride and urea, induce changes in the tertiary structure of proteins. We have presented results on such changes in four structurally unrelated proteins to show that such structural perturbations are common irrespective of their origin. Data representative of such structural changes are shown for the monomeric globular proteins such as horseradish peroxidase (HRP) from a plant, human serum albumin (HSA) and prothrombin from ovine blood serum, and for the membrane-associated, worm-like elongated protein, spectrin, from ovine erythrocytes. Structural alterations in these proteins were reflected in quenching studies of tryptophan fluorescence using the widely used quencher acrylamide. Stern-Volmer quenching constants measured in presence of the denaturants, even at concentrations below 100 mM, were higher than those measured in absence of the denaturants. Both steady-state and time-resolved fluorescence emission properties of tryptophan and of the extrinsic probe PRODAN were used for monitoring conformational changes in the proteins in presence of different low concentrations of the denaturants. These results are consistent with earlier studies from our laboratory indicating structural perturbations in proteins at the tertiary level, keeping their native-like secondary structure and their biological activity more or less intact.
Basak
S
S
Chemical Sciences Division, Saha Institute of Nuclear Physics, 1/AF Bidhannagar, Calcutta 700064, India. sbasak@hp1.saha.ernet.in
Debnath
D
D
Haque
E
E
Ray
S
S
Chakrabarti
A
A
D020106:Acrylamide / Q000494:pharmacology; D000818:Animals; D002942:Circular Dichroism; D004912:Erythrocytes / Q000737:chemistry; D006735:Horseradish Peroxidase / Q000737:chemistry*; D006801:Humans; D008956:Models, Chemical; D011489:Protein Denaturation; D011516:Prothrombin / Q000737:chemistry*; D012709:Serum Albumin / Q000737:chemistry*; D012756:Sheep; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014364:Tryptophan / Q000737:chemistry
D012709:Serum Albumin; D020106:Acrylamide; D014364:Tryptophan; D011516:Prothrombin; D006735:Horseradish Peroxidase