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Comparison of Backbone Dynamics of Oxidized and Reduced Putidaredoxin by 15N NMR Relaxation Measurements

Published

Author(s)

Fahriye N. Sari, Marcia J. Holden, M P. Mayhew, V L. Vilker, B Coxon

Abstract

The backbone dynamics of uniformly 15N-labeled reduced and oxidized putidaredoxin (Pdx) have been studied by 2D 15N NMR relaxation measurements. 15N T1 and T2 values and 1H-15N NOE s have been measured for the diamagnetic region of the protein. These data were analyzed by using a model free dynamics formalism to determine the generalized order parameters (S2), the effective correlation time for internal motions (τe) and 15N exchange broadening contributions (Rex) for each residue, as well as the overall correlation time (τm). Order parameters for the reduced Pdx are generally higher than for the oxidized Pdx, and there is increased mobility on the microsecond to millisecond time scale for the oxidized Pdx in comparison with the reduced Pdx. These results clearly indicate that the oxidized protein exhibits higher mobility than the reduced one, which is in agreement with the recently published redox-dependent dynamics studied by amide proton exchange. In addition, we observed very high T1/T2 ratios giving rise to a large Rex contribution for residues 33 and 34. Residue 34 is believed to be involved in the binding of Pdx to the cytochrome P450cam (CYP101). The differences in the backbone dynamics are discussed in relation to the oxidation states of Pdx and their impact on electron transfer.
Citation
Biochemistry
Volume
38
Issue
31

Keywords

backbone dynamics, correlation times, exchange broadening, nitrogen-15 NMR, NOE, order parameters, protein, putidaredoxin, relaxation times

Citation

Sari, F. , Holden, M. , Mayhew, M. , Vilker, V. and Coxon, B. (1999), Comparison of Backbone Dynamics of Oxidized and Reduced Putidaredoxin by <sup>15</sup>N NMR Relaxation Measurements, Biochemistry (Accessed February 26, 2024)
Created August 3, 1999, Updated February 19, 2017