The purified P68 virus was resuspended in Laemmli buffer and boiled for 3 min, and the proteins were separated by tricine gradient gel electrophoresis. All protein bands were cut from the gel and used for mass spectrometry (MS) analysis. In addition, P68 at a concentration of 2 mg/ml in phage buffer containing 1% SDS was boiled for 3 min and analyzed by MS. After destaining and washing, the proteins were subjected to trypsin digestion (sequencing grade, Promega). MALDI (matrix-assisted laser desorption/ionization)–MS and MS/MS analyses were performed on an Ultraextreme mass spectrometer (Bruker Daltonics, Bremen, Germany). Data processing and analysis were performed with the software FlexAnalysis 3.4 and MS BioTools (Bruker Daltonics). Mascot software (Matrix Science, London, UK) was used for sequence searches in exported MS/MS spectra against the National Center for Biotechnology Information database and a local database supplied with the expected protein sequences. The mass tolerance of peptides and MS/MS fragments for MS/MS ion searches were 50 parts per million and 0.5 Da, respectively. The oxidation of methionine and propionyl-amidation of cysteine as optional modifications and one enzyme miss-cleavage were set for all searches. Peptides with a statistically significant peptide score (P < 0.05) were considered.

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