N-15 NMR relaxation as a probe for helical intrinsic propensity: The case of the unfolded D2 domain of annexin I

The isolated D2 domain of annexin I is unable to adopt a tertiary fold but exhibits both native and non-native residual structures. It thus constitute s an attractive model for the investigation of dynamics of partially folded states in the context of protein folding and stability. N-15 relaxation pa rameters of the D2 domain have been acquired at three different magnetic fi elds, 500, 600 and 800 MHz. This enables the estimation of the contribution of conformational exchange to the relaxation parameters on the micro- to m illisecond time scale, thus providing a suitable data set for the descripti on of motions on the pico- and nanosecond time scale. The analysis of the s even spectral densities obtained (J(0), J(50 MHz), J(60 MHz), J(80 MHz), <J (500 MHz)>, <J(600 MHz)>, <J(800 MHz)>) provides complementary and meaningf ul results on the conformational features of the D2 domain structure previo usly depicted by chemical shift and NOE data. Especially, residual helix se gments exhibit distinct dynamical behaviors that are related to their intri nsic helical propensity. Beside the spectral density analysis, a series of models derived from the Lipari and Szabo model-free approach are investigat ed. Two models containing three parameters are able to reproduce equally we ll the experimental data within experimental errors but provide different v alues of order parameters and correlation times. The inability to find a un ique model to describe the data emphasizes the difficulty to use and interp ret the model-free parameters in the case of partially or fully unfolded pr oteins consisting of a wide range of interconverting conformers.