NMR SPECTROSCOPY OF ONE PEPTIDE SAMPLE

University Of Georgia, Athens GA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. NMR Spectroscopy The sample was first dissolved in a 90 % H2O/10 % D2O phosphate buffer at pH 3. The linewidth was too broad and only a few H alpha to NH crosspeaks could be detected. The sample was then lyophilized and redissolved in 600 uL 40 % EtOH/60 % H2O, which gave much better spectra. The sample started precipitating during acquisition;first an additional 50 uL H2O was added, which seemed to decrease the solubility, and so an additional 150 uL EtOH was added. The sample was warmed to ~40 [unreadable]C to dissolve as much as possible. The solution was transferred to a new NMR tube and maintained at 30 [unreadable]C to promote solubility. Spectra were recorded on a Varian Inova 800 MHz spectrometer at 30 [unreadable]C. Water suppression was achieved using 3919-Watergate sequences. A 60 ms TOCSY dataset was collected as a matrix of 600 X 3150 complex points, with acquisition times of 0.35 and 0.067 seconds in t2 and t1, respectively. A 150 ms NOESY dataset was collected as a matrix of 290 X 2048 complex points, with acquisition times of 0.23 and 0.032 seconds in t2 and t1. Data was processed with NMRPipe or Mestre-C with zero-filling in t1, application of Gaussian window functions in both dimensions prior to Fourier transform. Spectra were referenced to EtOH-d6 at 3.55 ppm.