EXPOSURE OF HYDROPHOBIC SURFACES ON THE CHAPERONIN GROEL OLIGOMER BY PROTONATION OR MODIFICATION OF HIS-401

Hydrophobic exposure on the chaperonin GroEL is increased 6-10-fold af ter the protein is treated with the His-reactive reagent diethyl pyroc arbonate (DEP), or the solution pH is lowered to 5.5. The induced hydr ophobic surfaces have the same 1,1'-bis(4-anilino)naphthalene-5,5'-dis ulfonic acid (bis-ANS) binding characteristics as unperturbed GroEL: a K-d congruent to 3.5 mu M, a maximum intensity at similar to 500 nm, and an average fluorescence lifetime of similar to 8.0 ns. The pK(alph a) for the pH-induced transition is 6.6, most likely attributable to t he only histidine in GroEL, His-401, located in the intermediate domai n. The modification of one histidine residue per monomer upon DEP trea tment is supported by the correlation between the change in the absorb ance at 242 nm for the N-carbethoxyhistidyl derivative and the increas e in bis-ANS fluorescence. GroEL at pH 5.5 is tetradecameric and can c apture urea-denatured rhodanese and release it as active enzyme. The G roEL rhodanese complex is more stable to dissociation by 2.25 M urea t han the complex formed at pH 7.8. We propose that His-401 is in a conf ormationally sensitive region such that protonation or modification ca n lead to increased expo sure of hydrophobic surfaces capable of bindi ng folding intermediates.