[13C]-constant-time [15N,1H]-TROSY-HNCA for sequential assignments of large proteins

The greatly improved sensitivity resulting from the use of TROSY during N-15 evolution and amide proton acquisition enables the recording of HNCA spectra of large proteins with constant-time C-13(alpha) evolution. In [C-13]-ct-[N-15,H-1]-TROSY-HNCA experiments with a H-2/C-13/N-15-labeled 110 kDa protein, 7,8-dihydroneopterin aldolase from Staphylococcus aureus, nearly all correlation peaks seen in the [N-15,H-1]-TROSY-HNCA spectrum were also detected. The improved resolution in the C-13 dimension then enabled a significant number of sequential assignments that could not be obtained with [N-15,H-1]-TROSY-HNCA without [C-13]-constant-time period.