Mix-and-match proteomics: using advanced iodoacetyl tandem mass tag multiplexing to investigate cysteine oxidation changes with respect to protein expression

Prakash, A. S., Kabli, A. M.F., Bulleid, N. and Burchmore, R. (2018) Mix-and-match proteomics: using advanced iodoacetyl tandem mass tag multiplexing to investigate cysteine oxidation changes with respect to protein expression. Analytical Chemistry, 90(24), pp. 14173-14180. (doi:10.1021/acs.analchem.8b02517) (PMID:30452864)

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Abstract

Cysteine redox state has been identified as one of the key biological influences behind protein structure and/or function. Altered protein redox state has been shown to cause significant physiological changes and can leave proteins with changed sensitivity to oxidative stress. Protein redox-state changes are recognized as an important mediator of disease, cellular abnormalities, and environmental changes, and therefore their characterization is of interest. Isotopic or isobaric labeling followed by sample multiplexing and analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) allows relative comparison of protein expression levels or of protein redox states between several samples. Combining analysis of protein expression level and redox state into one analysis would add an extra dimension and permit the normalization of protein redox changes with protein abundance. To achieve this, we have developed a quantitation workflow that uses commercially available cysteine-reactive tandem mass tags (iodoTMT) to differentially label cysteine residues, and we have applied it to two Leishmania mexicana cell lines that have previously shown different responses to oxidative stress. The individually labeled samples have been pooled in different combinations to create multiple sixplex samples in order to study the relationship between cysteine oxidation and overall protein expression, as well as providing information about protein oxidation levels in each cell line. The results highlight 11 proteins that are differentially expressed between the two cell lines and/or have significant redox changes. This advanced multiplexing method effectively demonstrates the flexibility of tandem mass tags and how they can be used to maximize the amount of information that can be acquired.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Bulleid, Professor Neil and Burchmore, Dr Richard and Kabli, Abdulbaset Mohammadmostafa F
Authors: Prakash, A. S., Kabli, A. M.F., Bulleid, N., and Burchmore, R.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Analytical Chemistry
Publisher:American Chemical Society
ISSN:0003-2700
ISSN (Online):1520-6882
Published Online:19 November 2018

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
172586Thiol Modification and Redox SignallingNeil BulleidMedical Research Council (MRC)MC_PC_15076Institute of Molecular, Cell & Systems Biology