This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Methods: NMR The samples (20 mg) were deuterium-exchanged by lyophilization from D2O (99.9 % D), dissolved in 0.7 mL D2O (99.96 % D), 100 ?L 10 mg/mL DSS in D2O (99.96 % D) was added, and the solution placed into a 5-mm NMR tube. One-dimensional proton NMR spectra were acquired on a Varian Inova-500 MHz instrument at 40 ?C. Spectral width was 4.7 kHz, transmitter frequency was set at -587 Hz, and 32 scans were collected in each experiment. The data were processed using the Mestre-C software using exponential multiplication (0.2 Hz). Spectra were referenced to the most upfield DSS signal (0.0 ppm). The spectra were baseline corrected with a third order Bernstein polynomial. The ratio of ?UA2S-H4 to ?UA-H4 was determined by integration of the signals at 5.97 ppm ( ?UA2S-H4) and 5.79 ppm (?UA-H4).
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