Post-Genomics and Skin Inflammation

Table 1

Oxidative PTMs of proteins and techniques for their study.

oxidative PTM	description
	gel-based approaches + MS protein identification	gel-free approaches
3-nitroTyrosine	WB detection with $\alpha$-nitroTyr antibodies [64–68]
4-HNE-adducts	WB detection with α-4-HNE antibodies [64, 69]	LC-ESI-MS/MS, as well as tryptic digestion, avidin column enrichment and MS/MS analysis following ${\text{N}}^{\prime }$-aminooxymethylcarbonylhydro-D-biotin labeling [70, 71]
Carbonylation	derivatization of protein carbonyls with 2,4-dinitrophenylhydrazine (DNPH) and WB detection of the resulting 2,4-dinitrophenylhydrazones (DNP) with $\alpha$-DNP antibodies; alternatively, biotin-hydrazide tagging and avidin-FITC staining [64, 66, 68, 69, 72, 73]	enrichment strategies with the hydrazide biotin-streptavidin methodology or Girard’s P reagent, coupled with LC-MS/MS and MALDI-MS/MS [66, 69–71]
Glycosylation	(i) lectin chemistry (due to a different specificity for certain carbohydrates, the identification of subclasses is allowed) [65]	(i) lectin affinity chromatography to isolate glycosylated proteins from complex mixtures prior to MS/MS [74]
	(ii) periodate/Schiff’s base chemistry to generate a general stain toward glycoproteins (Pro-Q Emerald staining) [65]	(ii) chemical trapping of N-glycosylated peptide prior to LC-MS/MS [74]
		(iii) chemoenzymatic or tagging via substrate strategies for O-glycolsylated peptided [74]
		(iv) COFRADIC [74]
Oxidation of -SH groups	various techniques to reveal specific -SH PTMs [66, 68, 73, 75]
	(i) oxidation, lack of labelling with specific reagents, such as biotinylated iodoacetamide, and WB detection with streptavidin	(i) isolation of cysteinyl peptides by biotinylation of Cys residues and affinity isolation (ICAT) [74]
	(ii) S-glutathionylation, metabolic labelling of the intracellular glutathione pool with ^35S-cysteine while inhibiting protein synthesis plus nonreducing electrophoresis, and autoradiography	(ii) S-nytrosilation, modified biotin switch method coupled with affinity isolation [74]
	(iii) S-nytrosilation, biotin switch method	(iii) COFRADIC [74]
	(iv) formation of disulphide bridges, diagonal 2-DE
	(v) sulfinic/sulfonic acid: detection through MS after standard trypsin digestion
Phosphorylation	(i) in gel protein staining with specific dies (Pro-Q Diamond) [65, 76]	(i) isolation of phosphopeptides by immobilized ion chromatography (IMAC) [74]
	(ii) WB with antibodies towards specific phosphorylated amino acids	(ii) segregation of phosphopeptides by strong cation exchange chromatography or titanium dioxide [74]
		(iii) various chemical reactions aiming at modifying the phopshorylated peptides prior to MS/MS [74]
		(iv) COFRADIC [74]
multiple PTMs	SEMSA during LC-ESI-MS/MS [77]