Evidence for extensive anisotropic local motions in a small enzyme using a new method to determine NMR cross-correlated relaxation rates in the absence of resolved scalar coupling

Abstract

Transverse 13CO-1HN (dipole-dipole)/13CO (CSA) cross-correlated relaxation rates were measured for the 13CO resonances of the protein ribonuclease Binase from Bacillus intermedius (12.3 kDa). This was carried out with a novel E.COSY-type triple-resonance experiment, which allows the measurement of cross-correlated transverse relaxation rate from multiplet effects in the absence of resolved scalar coupling. The 13CO-1HN (dipole-dipole)/13CO (CSA) cross-correlated relaxation rates were determined with an average precision of ±5% and cover a range of values between −1.5 and +0.6 Hz. The average (−0.44 Hz) is to be compared with the computed value of −0.83 Hz for this interaction. Mechanisms that potentially can cause the average to be smaller than the theoretical value and the unexpected large spread in observed values are discussed. It is suggested that large contributions to the variations are due to large amplitude local anisotropic motions.