The 2D {P} Spin-Echo-Difference Constant-Time [C, H]-HMQC Experiment for Simultaneous Determination of JH3′P and JC4′P in C-Labeled Nucleic Acids and Their Protein Complexes |
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| Authors: | Thomas Szyperski, C sar Fern ndez, Akira Ono, Kurt Wü thrich,Masatsune Kainosho |
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| Affiliation: | a Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule-Hönggerberg, CH-8093, Zürich, Switzerland;b Department of Chemistry, Faculty of Science, Tokyo Metropolitan University, 1-1 Minamiohsawa, Hachioji, 192-03, Japan |
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| Abstract: | A two-dimensional {31P} spin-echo-difference constant-time [13C, 1H]-HMQC experiment (2D {31P}-sedct-[13C, 1H]-HMQC) is introduced for measurements of 3JC4′P and 3JH3′P scalar couplings in large 13C-labeled nucleic acids and in DNA–protein complexes. This experiment makes use of the fact that 1H–13C multiple-quantum coherences in macromolecules relax more slowly than the corresponding 13C single-quantum coherences. 3JC4′P and 3JH3′P are related via Karplus-type functions with the phosphodiester torsion angles β and ε, respectively, and their experimental assessment therefore contributes to further improved quality of NMR solution structures. Data are presented for a uniformly 13C, 15N-labeled 14-base-pair DNA duplex, both free in solution and in a 17-kDa protein–DNA complex. |
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| Keywords: | 3JC4′ P and 3JH3′ P scalar couplings protein– nucleic acid complexes multiple-quantum coherence NMR structure determination 13C labeling |
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