Assignments of carbon NMR resonances for microcrystalline ubiquitin. Academic Article uri icon

abstract

  • Solid-state NMR 2D spectroscopy was used to correlate carbon backbone and side-chain chemical shifts for uniformly (13)C,(15)N-enriched microcrystalline ubiquitin. High applied field strengths, 800 MHz for protons, moderate proton decoupling fields, 80-100 kHz, and high magic angle sample spinning frequencies, 20 kHz, were used to narrow the most of the carbon line widths to 0.5-0.8 ppm. Homonuclear magnetization transfer was effected by matching the proton RF field to the spinning frequency, the so-called dipolar-assisted rotational resonance (DARR) (Takegoshi, K.; Nakamura, S.; Terao, T. Chem. Phys. Lett. 2001, 344, 631-637), and a mixing time of 20 ms was used to maximize the intensity of one-bond transfers between carbon atoms. This polarization transfer sequence resulted in roughly 14% transfer efficiencies for directly bonded carbon pairs and 4% transfer efficiencies for carbons separated by a third carbon. With this simple procedure, the majority of the one-bond correlations was observed with moderate transfer efficiencies, and many two-bond correlations were also observed with weaker intensities. Spin systems could be identified for more than half of the amino acid side chains, and site-specific assignments were readily possible via comparison with 400 MHz (15)N-(13)C-(13)C correlation spectroscopy (described separately).

published proceedings

  • J Am Chem Soc

altmetric score

  • 3

author list (cited authors)

  • Igumenova, T. I., McDermott, A. E., Zilm, K. W., Martin, R. W., Paulson, E. K., & Wand, A. J.

citation count

  • 185

complete list of authors

  • Igumenova, Tatyana I||McDermott, Ann E||Zilm, Kurt W||Martin, Rachel W||Paulson, Eric K||Wand, A Joshua

publication date

  • June 2004