A JCH-Modulated 2D (HACACO)NH Pulse Scheme for Quantitative Measurement of 13Cα–1Hα Couplings in 15N, 13C-Labeled Proteins

✍ Scribed by T.Kevin Hitchens; Scott A. McCallum; Gordon S. Rule

• Publisher: Elsevier Science
• Year: 1999
• Tongue: English
• Weight: 73 KB
• Volume: 140
• Category: Article
• ISSN: 1090-7807
• DOI: 10.1006/jmre.1999.1847

✦ Synopsis

A triple-resonance pulse sequence is presented for the quantitative measurement of 1 H ␣ -13 C ␣ single-bond couplings in 15 N, 13 C uniformly labeled proteins. This 1 J CH -modulated (HACACO)NH experiment yields 1 H N -15 N-correlated 2D spectra in which the amplitude of each peak is modulated by the 1 H ␣ -13 C ␣ J coupling of the preceding residue, (i ؊ 1). The experiment is demonstrated on a 1.0 mM sample of Rho130, the 15-kDa RNA binding domain of E. coli Rho factor. The average error in the measured coupling constants was less than 0.8%. This sequence allows the measurement of the 1 J CH couplings from a proton-nitrogen HSQC without the need for assigning the H ␣ and C ␣ resonances.