UNFOLDING AND AGGREGATION-ASSOCIATED CHANGES IN THE SECONDARY STRUCTURE OF D-GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE DURING DENATURATION BY GUANIDINE-HYDROCHLORIDE AS MONITORED BY FTIR

The secondary structure of native D-glyceraldehyde-3-phosphate dehydro genase was compared with its partially folded intermediate and aggrega ted states obtained during guanidine hydrochloride (GdnHCl) denaturati on using transmission Fourier transform infrared (FTIR) and micro-FTIR measurements. The changes in the secondary structures indicated a par tially folded intermediate formed in 0.1 M GdnHCl solution without vis ible aggregation. Increasing the GdnHCl concentration resulted in aggr egation of the enzyme along with changes in the secondary structure. A lthough similar relative amounts of the secondary structure were found in the aggregated and native states of the enzyme, the temporal varia tion of the secondary structure revealed a difference in the beta-shee t structure between the aggregated and native states, suggesting that the aggregation resulted from further unfolding of the enzyme. In addi tion, FTIR data suggest that such aggregates are most likely mediated by specific intermolecular interactions and that the predominant drivi ng force involved in aggregation may be a hydrophobic interaction betw een exposed surfaces of partially folded intermediates. (C) 1997 John Wiley & Sons, Inc.