Project description:Global proteome study of protein extracts of purified Tupavirus, using a bidimensionnal nanoLC fractionnation combined to a Synapt G2 Si HDMSe monitoring enabling ion mobility
Project description:Global proteome study of two protein extracts of purified Faustovirus, using a bi dimensionnal nanoLC fractionnation combined to a Synapt G2 Si HDMSe monitoring enabling ion mobility
Project description:Comparison among the following data independent acquisition modes provided on the Waters Synapt G2-S instrument: MSE, MSE with Ion Mobility (HDMSE), and MSE with Ion Mobility and drift time specific collision energies (UDMSE). The following on-column loads and LC gradient lengths have been used for the DDA data (1-20130710-63647): * 200 ng, 90 minutes gradient * 300 ng, 180 minutes gradient * 1000 ng, 180 minutes gradient. DDA data has been processed by using MaxQuant (v.1.3.0.5) software, searching against a organism specific (Homo Sapiens) Uniprot/Swissprot database. FDR has been controlled by using a pseudo-reverse (KR positions kept) database at both protein and peptide levels to 1%. Proteins with less than two peptides (minimum of 6 amino acids length, up to 2 missed cleavages and 3 variable modifications allowed. Variable identifications were : methionine oxidation, fixed modifications were : cysteine carbamidomethylation ) identified have been discarded. A match between runs of a 2 minutes windows among the three technical replicates has been performed.
Project description:These are supporting RAW files for 'Detergent-free simultaneous sample preparation method for proteomics and metabolomics' SiTrap method manuscript.
Frozen renal tissue from three matched clear cell renal carcinoma (G2 pT3a, G2 pT1b, G1 pT2) /adjacent normal sample pairs were obtained from The Leeds Multidisciplinary Research Tissue Bank. Approximately 1 cm2 sections with a thickness of 10 um were lysed and processed according to SiTrap protocol using SiTrap cellulose tips. The SiTrap load was normalized by protein concentration. 50 ug of protein was loaded. The collected flow-through fraction, devoid of proteins, was dried down for targeted metabolomics analysis. The captured protein fraction was reduced and alkylated in situ and digested with trypsin according to SiTrap protocol. The resulting peptides were concentrated for proteomics analysis.