Candida Proteome Profiling by LC-MS/MS

LC-MS/MS analysis was carried out for peptide samples using an EASY NanoLC 1000 system (ThermoFisher Scientific) coupled on-line with an LTQ-Orbitrap XL MS (ThermoFisher Scientific) as described [84 (link)]. Each cycle of an MS/MS experiment includes one MS scan in FT mode (350–1400 m/z, resolution of 60,000 at m/z 400) followed by data-dependent MS2 scans in the LTQ with normalized collision energy at 35% on the top ten peaks. Raw data were searched against a database consisting of Candida proteins (PA.orf_trans_all_assembly_21_2009_0306_v2 with a total of 6243 protein entries) using MaxQUANT. The mass tolerance for parent ions and fragment ions were set as ± 20 ppm and 0.5 Da, respectively. Trypsin was set as the enzyme, and a maximum of two missed cleavages were allowed. Protein N-terminal acetylation and methionine oxidation were selected as variable modifications. Protein and PSM FDR was set as 0.01. LFQ and iBAQ protein quantitation were calculated using a minimum ratio count of 2 peptides [46 (link), 47 (link)]. All raw data was deposited and can be accessed at ftp://MSV000081642@massive.ucsd.edu (username, if prompted: MSV000081642; password: SA10242017UCI).

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Alkafeef S.S., Yu C., Huang L, & Liu H. (2018). Wor1 establishes opaque cell fate through inhibition of the general co-repressor Tup1 in Candida albicans. PLoS Genetics, 14(1), e1007176.

Publication 2018

EASY NanoLC 1000 system LTQ-Orbitrap XL MS

Acetylation Candida Cleavages Enzyme Ions Methionine Ms ms Parent Peptides Protein Protein a Protein n Scans Tolerance Trypsin

Corresponding Organization : University of California, Irvine

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Variable analysis

independent variables

LC-MS/MS analysis
Database searching using MaxQUANT

dependent variables

Peptide identification and quantification

control variables

Trypsin enzyme digestion
Protein N-terminal acetylation and methionine oxidation as variable modifications
Protein and PSM FDR set at 0.01
Minimum ratio count of 2 peptides for LFQ and iBAQ protein quantitation

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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