Bacterial chaperonin GroEL with a molecular mass of 800 kDa was studied by (13)C NMR spectroscopy. Carbonyl carbons of GroEL were labeled with (13)C in an amino acid specific manner in order to reduce the number of signals to be observed in the spectrum. Combination of selective labeling and site-directed mutagenesis enabled us to establish the sequence specific assignment of the (13)C resonances from GroEL. ADP-binding induced a chemical shift change of Tyr478 in the equatorial domain and His401 in the intermediate domain, but little of Tyr203 in the apical domain. Upon complex formation with co-chaperonin GroES in the presence of ADP, Tyr478 exhibits two peaks that would originate from the cis and trans rings of the asymmetric GroEL-GroES complex. Comparison between the line width of the GroEL resonances and those from GroES in complex with GroEL revealed broadening disproportionate to the size of GroEL, implying the existence of conformational fluctuations which may be pertinent to the chaperone activity. Based on these results, we concluded that (13)C NMR observation in combination with selective labeling and site-directed mutagenesis can be utilized for probing the conformational change and dynamics of the extremely large molecules that are inaccessible with current NMR methods.