Understanding groel function by means of sulfhydryl labeling

Confot mat ion al changes of GroKL complexée! with adr-nine nucleotides. Mg"1"", or rhodanese wore- studied by examining the labeling patterns of a sulfhvdryl directed probe, fluorescein-ô- rnaleimide. GroEI, contains three sulfhydrvis: Cysl.'JH. Cys45S, and C'ys519. Our data indicate that rearrangements are o< earring in the structure of GroKI. upon tho binding of these various ligands I'nder all conditions. Cys138 was labeled whereas (Vs519 was never labeled under any of the conditions. The efficiency of labeling Cys45K varied greatlv depending on which ligand was used. Cys458 is located at the interface be tween the two GroKL heptamers in the equatorial domain. Cys-î58 is labeled moderately under normal buffer conditions but is labeled extensively when GroKL is exposed to Mg+2 or rhodanese. However, when GroEI, is exposed to MgADP, MgATP, or MgAMP-PNP, the labeling is strongly diminished. The differences of labeling efficiency may be the result of dynamic conformational changes occurring when ligands as well as substrate protein are bound. Inter estingly. our data also indicate that Mg+2 alone changes the conformation of GroEI.