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  • Title: Sensitivity improvement for correlations involving arginine side-chain Nepsilon/Hepsilon resonances in multi-dimensional NMR experiments using broadband 15N 180 degrees pulses.
    Author: Iwahara J, Clore GM.
    Journal: J Biomol NMR; 2006 Dec; 36(4):251-7. PubMed ID: 17036160.
    Abstract:
    Due to practical limitations in available 15N rf field strength, imperfections in 15N 180 degrees pulses arising from off-resonance effects can result in significant sensitivity loss, even if the chemical shift offset is relatively small. Indeed, in multi-dimensional NMR experiments optimized for protein backbone amide groups, cross-peaks arising from the Arg guanidino 15Nepsilon (approximately 85 ppm) are highly attenuated by the presence of multiple INEPT transfer steps. To improve the sensitivity for correlations involving Arg Nepsilon-Hepsilon groups, we have incorporated 15N broadband 180 degrees pulses into 3D 15N-separated NOE-HSQC and HNCACB experiments. Two 15N-WURST pulses incorporated at the INEPT transfer steps of the 3D 15N-separated NOE-HSQC pulse sequence resulted in a approximately 1.5-fold increase in sensitivity for the Arg Nepsilon-Hepsilon signals at 800 MHz. For the 3D HNCACB experiment, five 15N Abramovich-Vega pulses were incorporated for broadband inversion and refocusing, and the sensitivity of Arg1Hepsilon-15Nepsilon-13Cgamma/13Cdelta correlation peaks was enhanced by a factor of approximately 1.7 at 500 MHz. These experiments eliminate the necessity for additional experiments to assign Arg 1Hepsilon and 15Nepsilon resonances. In addition, the increased sensitivity afforded for the detection of NOE cross-peaks involving correlations with the 15Nepsilon/1Hepsilon of Arg in 3D 15N-separated NOE experiments should prove to be very useful for structural analysis of interactions involving Arg side-chains.
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