LC-MS/MS analysis identified CgCrzA phosphorylation sites by CgCrzA.

To identify the phosphorylation sites of CgCrzA, total proteins were extracted from the WT/CgCrzA-GFP and DCgmkk1/CgCrzA-GFP transformants, and subjected to purification steps described previously. Purified proteins were separated via 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Gel bands corresponding to CgCrzA were excised and analyzed using a Thermo Q-Exactive high-resolution mass spectrometer (Thermo Scientific, Waltham, MA). Phosphorylated sites were identified by MS/MS spectra data with Mascot Distiller (Matrix Science; version 2.4).