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Carbamidomethylation of Cys (C) residues were set as fixed modifications, and oxidation of Met and acetylation of N-terminal G1 and K2 residues were set as variable modifications.
Other parameters include methylthiol modification of cysteine, iTRAQ labels at the peptide N-terminus and Lysine residues as static modifications and oxidation of methionine as variable modification.
For the global proteomic dataset, carbamidomethylation of cysteine and iTRAQ 4-plex modification of peptide N-termini were set as fixed modifications, and oxidation of methionine and iTRAQ 4-plex modification of Lys were set as variable modifications.
Further settings used were the following: trypsin with 1 missed cleavage; carbamidomethylation on cysteine and iTRAQ-8plex on lysine and N-terminal as fixed modifications; and oxidation of methionine as variable modification.
The parameters used for data analysis included trypsin as a protease (allowed one missed cleavage), ITRAQ labeling at N-terminus and lysine residues, and cysteine modification by methyl methane thiosulfonate as fixed modifications and oxidation of methionine as a variable modification.
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To experimentally differentiate the contributions of oxidative processes that could lead to protein modifications and oxidation-independent proteasome inhibitor activity of heme, we performed a screening of different metal porphyrins with the aim of identifying non-iron porphyrins with significant proteasome inhibitor activity, but absent oxidative activity.
Carbamidomethylation of cysteine was set as fixed modification and oxidation of methionines as variable modification.
Carbamidomethylcysteine was selected as a fixed modification and oxidation of methionine as a variable modification.
Carbamidomethylation (C) was set as a fixed modification, and oxidation (M), phosphorylation (S, T and Y) and deamidation (N) were set as variable modifications.
The following parameters were used for the database search: MS and MS/MS accuracies were set to <0.6Da, trypsin/P as an enzyme, missed cleavages 1, carbamidomethylation of cysteine as fixed modification, and oxidation of methionine as a variable modification.
CID spectra were interpreted by Phenyx software (Genebio SA, Geneva, Switzerland) using carboamidomethylation of cysteine as a fixed modification, and oxidation of methionine and EPG on glutamic acid as variable modification.
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