For ²H-labeled proteins, it is necessary to use a slighlty modifed 3D HNCA experiment:

Deuterium decoupling is applied during the ¹³C-evolution constant-time t₁ period when the magnetization is on the alpha carbons.

The broadband ¹H-decoupling is interrupted during this time period and proton-carbon couplings are refocused by a 180o ¹H-pulse.

Create a new dataset (new) and read the standard BRUKER parameter set (rpar) to record a sensitivity-enhanced 3D HNCA experiment with ²H-decoupling by typing rpar HNCAGP2H3D all. The pulse program hncagp2h3d can be visualized in the PulsProg section or with the edcpul command).

The major modifications, with respect to the original experiment, are the following:

• The nucleus on f4 channel is defined as NUC4=2H
• o4p=3.0ppm, centered to the ²H aliphatic region.
• Deuteron decoupling is achieved by applying cpdgrp4=waltz16, at power level pl17, using a decoupling pulse length about pcpd4=300us.