LC-MS/MS analysis
HERC5-Flag, along with WT or MUT OspC1, overexpressed in 293T cells was
immunoprecipitated with anti-Flag M1 agarose affinity gel
(Sigma-Aldrich, A4596-5ml) 24 h after transfection and then subjected to
SDS-PAGE separation and Coomassie Blue staining. Target HERC5-Flag bands
were taken for in-gel digestion with trypsin according to a standard
protocol, followed by Liquid chromatography-tandem mass spectrometry
(LC-MS/MS) analysis on Thermo ScientificTM Orbitrap
FusionTM coupled with UltiMate3000. The MS/MS spectra
of modified peptides were acquired by searching the raw data with Mascot
(version 2.3.02, Matrix Science).