The DQ-HN{CACB} and DQ-HN(CO){CACB} sequences with evolution of double quantum C α–C β coherences

Abstract

The new variant of known HNCACB and HN(CO)CACB techniques is proposed that employs excitation and evolution of double quantum C α–C β coherences. The most important features of the new method are: increased signal dispersion, lack of splittings due to 1 J(C α–C β) spin–spin couplings, and absence of accidental cancellations of positive and negative signals. The acquisition of both DQ-HN{CACB} and DQ-HN(CO){CACB} techniques enables sequential assignment of protein backbone, using only C α–C β DQ-frequencies. The determination of all C α and C β chemical shifts requires, however, a comparison with HN(CO)CA or HNCA spectra. Examples of applications of the DQ-HN{CACB} and DQ-HN(CO){CACB} experiments are presented, employing the 2D Reduced Dimensionality approach for 13C, 15N-labeled ubiquitin, and the 3D acquisition for 13C, 15N-double labeled Ca 2+-binding bovine S100A1 protein in the apo state (21 kDa) with overall correlation time of 8.1 ns.