Measurement of carbonyl chemical shifts of excited protein states by relaxation dispersion NMR spectroscopy: comparison between uniformly and selectively 13C labeled samples

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Measurement of carbonyl chemical shifts of excited protein states by relaxation dispersion NMR spectroscopy: comparison between uniformly and selectively 13C labeled samples
Patrik Lundström, D. Flemming Hansen and Lewis E. Kay
Journal of Biomolecular NMR; 2008; 42(1); pp 35 - 47

Abstract: Carr–Purcell–Meiboom–Gill (CPMG) relaxation dispersion nuclear magnetic resonance (NMR) spectroscopy has emerged as a powerful method for quantifying chemical shifts of excited protein states. For many applications of the technique that involve the measurement of relaxation rates of carbon magnetization it is necessary to prepare samples with isolated 13C spins so that experiments do not suffer from magnetization transfer between coupled carbon spins that would otherwise occur during the CPMG pulse train. In the case of 13CO experiments however the large separation between 13CO and 13Cα chemical shifts offers hope that robust 13CO dispersion profiles can be recorded on uniformly 13C labeled samples, leading to the extraction of accurate 13CO chemical shifts of the invisible, excited state. Here we compare such chemical shifts recorded on samples that are selectively labeled, prepared using [1-13C]-pyruvate and NaH13CO3, or uniformly labeled, generated from 13C-glucose. Very similar 13CO chemical shifts are obtained from analysis of CPMG experiments recorded on both samples, and comparison with chemical shifts measured using a second approach establishes that the shifts measured from relaxation dispersion are very accurate.