Refolding of thioredoxin reductase assisted by groEL and PDI

Thioredoxin reductase was unfolded in 2 M guanidine hydrochloride as reveal ed by fluorescence and CD spectroscopy. Spontaneous refolding of denatured species resulted in low recovery of 10% catalytic activity after 4 h incuba tion at 25 degrees C. Addition of groEL or protein disulfide isomerase to t he renaturation buffer accelerated the rate of recovery of catalytic activi ty to a level of 35 and 15%, respectively. Fluorescence spectroscopy has be en used to investigate the interaction of groEL and protein disulfide isome rase with denatured thioredoxin reductase tagged with a fluorescent probe. The fluorescence emitted by the denatured protein was quenched upon binding to either groEL or protein disulfide isomerase. It is suggested that encap sulation of the protein substrate by the chaperone plays an important role in the process of folding by facilitating the formation of correctly folded species. (C) 1999 Academic Press.