Mass Spectrometry Proteomics Protocol

The full-length MS analyses were performed by the University of Arkansas Statewide Mass Spectrometry Facility. Purified proteins were dialyzed with pure water, diluted to 0.05 mg mL⁻¹, and analyzed by a Bruker UltraflexII TOF-TOF with a MALDI ionization source. The laser was an Azura Laser AG diode pumped solid state laser (Nd:YAG, 355nm). The matrix used was a saturated solution of sinipinic acid in water/acetonitrile. Spectra were obtained in the positive ion, linear mode. The LC-MS/MS analyses were performed by Yale University Keck Proteomics Facility. The protocol was the same as the previous study.^{56 (link)} Proteins were digested by trypsin with a standard in-gel digestion protocol, and analyzed by LC-MS/MS on an LTQ Orbitrap XL equipped with a nanoACQUITY UPLC system. The Mascot search algorithm was used to search for the appropriate noncanonical substitution (Matrix Science, Boston, MA).