Simultaneous measurement of 1H–15N and Methyl 1Hm–13Cm residual dipolar couplings in large proteins

A two-dimensional TROSY-based SIM-13Cm–1Hm/1H–15N NMR experiment for simultaneous measurements of methyl 1DCH and backbone amide 1DNH residual dipolar couplings (RDC) in {U-[15N,2H]; Ileδ1-[13CH3]; Leu,Val-[13CH3/12CD3]}-labeled samples of large proteins is described. Significant variation in the alignment tensor of the 82-kDa enzyme Malate synthase G is observed as a function of only slight changes in experimental conditions. The SIM-13Cm–1Hm/1H–15N data sets provide convenient means of establishing the alignment tensor characteristics via the measurement of 1DNH RDCs in the same protein sample.