The 3D H(CC)(CO)NH experiment is specifically designed to correlate the ¹H and ¹⁵N amide resonances of one residue with ¹HA and all other ¹H side-chain resonances of its preceding residue via the intervening ¹³CO and ¹³C_aliphatic spins by means of the ¹J(NH), ¹J(N,CO), ¹J(CA,CO) ¹J(C,C),and ¹J(H,C) coupling constants. This experiments is closely related to the 3D (H)CC(CO)NH experiment.

Implementation on AVANCE spectrometers equipped with a third channel. Improved versions using pulsed field gradients (PFGs) are also available and, therefore, in such cases gradient technology is required.

The experiment is applied on ¹⁵N,¹³C-labeled proteins. Because the amide (NH) protons are involved, the  (H)CC(CO)NH experiment must be recorded in H₂O.

The 3D H(CC)(CO)NH pulse sequence ( 92JACS10974 , 93JMRB114-101 , 93JMRB206-101 , 93JB225 ) consists of the following steps:

1. ¹H chemical shift evolution during a t₁ period followed by a transfer to all aliphatic ¹³C via ¹J(CH) using an INEPT pulse sequence.
2. ¹³C-¹³C DIPSI-2 pulse train transfers magnetization from all side chain ¹³C resonances to ¹³CA via ¹J(CC).
3. Fixed evolution delay to achieve antiphase ¹³CA magnetization with respect to ¹³CO via ¹J(CA,CO) followed by  magnetization transfer to ¹³CO by simultaneous 90º ¹³C and ¹³CO pulses.
4. Fixed evolution delay to achieve antiphase ¹³CO magnetization with respect to ¹⁵N via ¹J(N,CO) followed by  magnetization transfer to ¹⁵N by simultaneous 90º ¹⁵N and ¹³CO pulses.
5. ¹⁵N chemical shift evolution during the constant-time t₂ period. Finally, magnetization is transferred back to the NH protons by a retro-INEPT pulse train and proton acquisition is performed under ¹⁵N decoupling.

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Several improved versions have been proposed ( 98JB191 and 99PROG93) even for large duterated proteins ( 99JB227).

Version using 1H-13C cross-polarization transfer and 13C-13C TOCSY transfer .

Version using WATERGATE

A 3D TROSY-based version has been proposed for a 2H-labeled membrane protein ( 02JB289)

In addition, related 2D H(CC)(CO)(N)H (  96JB161 ) 4D HCC(CO)NH experiments have been also reported (   92FEBS413 and   93JB349 ).

A related 3D H(CC)NH experiment can be also used to obtain sequential conectivities ( 99JB227)

The 3D H(CC)(CO)NH experiment can be recorded in automation mode.

The H(CC)(CO)NH experiment affords a 3D spectrum in which ¹H (sidechain), ¹⁵N and ¹HN chemical shifts are displayed in three independent dimensions. Sequential connectivities are due to ¹J(NH) +  ¹J(N,CO) +  ¹J(CA,CO) +  n*¹J(C,C) + ¹J(C,H).

See list of 3D triple-resonance NMR experiments for doubly-labeled proteins.