FOLLOWING PROTEIN-FOLDING IN REAL-TIME USING NMR-SPECTROSCOPY

The refolding of apo bovine a-lactalbumin has been monitored in real t ime by NMR spectroscopy following rapid in situ dilution of a chemical ly denatured state. By examining individual resonances in the time-res olved NMR spectra, the native state has been shown to emerge in a coop erative manner from an intermediate formed in the dead-time of the exp eriments. The kinetics of folding to the native state are closely simi lar to those observed by stopped-flow fluorescence and near-UV circula r dichroism. The NMR spectrum of the transient intermediate resembles closely that of the well characterized stable molten globule state for med at low pH. The results suggest that NMR can play a key role in des cribing at an atomic level the structural transitions occurring during protein folding.