Recently proposed triple-resonance experiments (1-14) make it possible to obtain complete backbone assignments of isotopically enriched proteins without recourse to NOE information. A main practical problem with the triple-resonance assignment approach is the necessity to record a substantial number of 3D spectra, each requiring typically at least one or two days of measuring time. A 4D triple-resonance approach has been proposed to remove the need for recording more than just two 4D experiments for completing the backbone assignments ( 11). However, if there is overlap in the ‘H,-13C, region of the 2D spectrum, these and other (12) 4D experiments also do not offer unique connections between sequential amides. In cases of such overlap, an experiment that relays magnetization from the amide to the intraresidue carbonyl can resolve the ambiguity caused by the C,-H, degeneracy ( 13). Alternatively, an experiment named CBCA(CO)NH, which correlates amide signals with the C, and Cs reson.ances of the preceding residue by using relay via the intervening i3C0 nucleus, provides information on the type of amino acid preceding each amide and thus frequently resolves the ambiguity caused by C,-H, degeneracy (14). Here we describe a similar experiment, named CBCANH, which correlates the amide ‘H and “N signals with ‘both the intraresidue C, and C, signals and with the C, and C, of the preceding residue. For smalland medium-sized proteins, up to -2OkDa, this 3D experiment alone can yield virtually complete assignments of the backbone and the C, resonances. The pulse scheme for the CBCANH experiment is sketched in Fig. 1. Before describing the experiment in more detail, we first outline its general mechanism. An INEPT transfer enhances polarization of both C, and CB spins. Chemical shifts of both C, and CB are encoded during the constant-time evolution period, 2TAB ( 7, 8, 10, Z4). When the 90 ’ 13C pulse is applied, at time b in Fig. 1, CB magnetization is transferred to C,, but at the same time a fraction of C, magnetization remains on C,. Subsequently, C, magnetization is transferred to its intraresidue 15N and to the 15N of the next residue, via the ’ JColN and 2 JcaN couplings, respectively. Following a constant-time 15N evolution period, the “N magnetization is transferred to the amide protons with a reverse INEPT sequence, prior to detection of the HN signals.