Backbone dynamics of the n-terminal domain in E-coli DnaJ determined by N-15- and (CO)-C-13-relaxation measurements

The backbone dynamics of the N-terminal domain of the chaperone protein Esc herichia coli DnaJ have been investigated using steady-state H-1-N-15 NOEs, N-15 T-1, T-2, and T-1 rho, relaxation times, steady-state C-13(alpha)-(CO )-C-13 NOEs, and (CO)-C-13 T-1 relaxation times. Two recombinant constructs of the N-terminal domain of DnaJ have been studied. One, DnaJ(1-78), conta ins the most conserved "J-domain" of DnaJ, and the other, DnaJ(1-104), incl udes a glycine/phenylalanine rich region ("G/F" region) in addition to the "J-domain". DnaJ(1-78) is not capable of stimulating ATP hydrolysis by DnaK , despite the fact that all currently identified sites responsible for DnaJ -DnaK interaction are located in this region. DnaJ(1-104), on the other han d, retains nearly the full ATPase stimulatory activity of full length DnaJ. Recently, a structural analysis of these two molecules was presented in an effort to elucidate the origin of their functional differences [Huang, K,, Flanagan, J. M., and Prestegard, J. H. (1999) Protein Science 8, 203-214]. Herein, an analysis of dynamic properties is presented in a similar effort . A generalized model-free approach with a full treatment of the anisotropi c overall rotation of the proteins is used in the analysis of measured rela xation parameters. Our results show that internal motions on pico- to nanos econd time scales in the backbone of DnaJ(1-78) are reduced on the inclusio n of the "G/F" region, while conformational exchange on micro- to milliseco nd time scales increases. We speculate that the enhanced flexibility of res idues on the slow time scale upon the inclusion of the "G/F" region could b e relevant to the ATPase stimulatory activity of DnaJ if an "induced-fit" m echanism applies to DnaJ-DnaK interactions.