Entrapping intermediates of thermal aggregation in alpha-helical proteins with low concentration of guanidine hydrochloride.
Aggregation of proteins is a problem with serious medical implications and economic importance. To develop strategies for preventing aggregation, the mechanism(s) and pathways by which proteins aggregate must be characterized. In this study, the thermally induced aggregation processes of three alpha-helix proteins (myoglobin, cytochrome c, and lysozyme) in the presence and absence of 1.0 m guanidine hydrochloride (GdnHCl) were investigated by means of infrared spectroscopy. In the absence of GdnHCl, intensities of the alpha-helix bands (approximately 1656 cm(-1)) decrease as a function of temperature at above 50 degrees C. With myoglobin and cytochrome c, the loss of helix bands was accompanied by the appearance of two new bands at 1694 and 1623 cm(-1), indicative of the formation of intermolecular beta-sheet aggregates. For lysozyme, bands indicative of intermolecular beta-sheet aggregates did not appear in any significant intensity. In the presence of 1.0 m GdnHCl, two major intermediate states rich in 3(10)-helix (represented by the band at 1663 cm(-1)) and beta-turn structure (represented by the band at 1667 cm(-1)), respectively, were observed. These findings demonstrated that IR spectroscopic studies of protein aggregation using a combination of thermal and chemical denaturing factors could provide a means to populate and characterize aggregation intermediates.
Dong
A
A
Department of Chemistry and Biochemistry, University of Northern Colorado, Greeley, Colorado 80639, USA. adong@unco.edu
Randolph
T W
TW
Carpenter
J F
JF
D000818:Animals; D002645:Chickens; D003574:Cytochrome c Group / Q000737:chemistry*; D019791:Guanidine / Q000494:pharmacology*; D006736:Horses; D006358:Hot Temperature; D009113:Muramidase / Q000737:chemistry*; D009211:Myoglobin / Q000737:chemistry*; D017433:Protein Structure, Secondary / Q000187:drug effects; D013055:Spectrophotometry, Infrared / Q000379:methods; D013816:Thermodynamics
D003574:Cytochrome c Group; D009211:Myoglobin; D009113:Muramidase; D019791:Guanidine