The use of heteronuclear cross-polarization for backbone assignment of (2)H-, (15)N- and (13)C-labeled proteins: A pulse scheme for triple-resonance 4D correlation of sequential amide protons and (15)N

M Shirakawa; M Wälchli; M Shimizu; Y Kyogoku

doi:10.1007/BF00211761

The use of heteronuclear cross-polarization for backbone assignment of (2)H-, (15)N- and (13)C-labeled proteins: A pulse scheme for triple-resonance 4D correlation of sequential amide protons and (15)N

J Biomol NMR. 1995 Apr;5(3):323-6. doi: 10.1007/BF00211761.

Authors

M Shirakawa¹, M Wälchli, M Shimizu, Y Kyogoku

Affiliation

¹ Institute for Protein Research, Osaka University, Suita, Osaka 565, Japan.

PMID: 22911505
DOI: 10.1007/BF00211761

Abstract

A new four-dimensional pulse scheme is described for the main-chain assignment of proteins by means of the J connectivity of the amide proton and nitrogen resonances of adjacent residues. Since the new experiment, 4D CP-HN(COCA)NH, involves heteronuclear cross-polarization for magnetization transfer from (13)C=O to (15)N via (13)C(α), a relatively strong WALTZ-16 decoupling rf field is applied to (13)C(α) during magnetization transfer. Consequently, (13)C(α) is effectively decoupled from its attached (2)H in the case of deuterated proteins, in the absence of a decoupling rf field for (2)H. This efficiently improves the sensitivity of the experiment through (13)C line narrowing. The experiment was performed on a randomly 60% deuterated protein, and the sensitivity of the final 4D spectrum was found to be excellent.