Quantitative Peptidomics with Five-plex Reductive Methylation labels

Autor(es)

Tashima, Alexandre K. ^[1] ; Fricker, Lloyd D. ^{[2, 3]}

Afiliação do(s) autor(es)

^[1] Univ Fed Sao Paulo, Escola Paulista Med, Dept Biochem, BR-04023901 Sao Paulo, SP - Brazil

^[2] Albert Einstein Coll Med, Dept Neurosci, Bronx, NY 10461 - USA

^[3] Albert Einstein Coll Med, Dept Mol Pharmacol, Bronx, NY 10461 - USA

• Tipo de documento: Artigo Científico
• Fonte: JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY; v. 29, n. 5, p. 866-878, MAY 2018.
• Citações Web of Science: 0

Resumo

Quantitative peptidomics and proteomics often use chemical tags to covalently modify peptides with reagents that differ in the number of stable isotopes, allowing for quantitation of the relative peptide levels in the original sample based on the peak height of each isotopic form. Different chemical reagents have been used as tags for quantitative peptidomics and proteomics, and all have strengths and weaknesses. One of the simplest approaches uses formaldehyde and sodium cyanoborohydride to methylate amines, converting primary and secondary amines into tertiary amines. Up to five different isotopic forms can be generated, depending on the isotopic forms of formaldehyde and cyanoborohydride reagents, allowing for five-plex quantitation. However, the mass difference between each of these forms is only 1 Da per methyl group incorporated into the peptide, and for many peptides there is substantial overlap from the natural abundance of C-13 and other isotopes. In this study, we calculated the contribution from the natural isotopes for 26 native peptides and derived equations to correct the peak intensities. These equations were applied to data from a study using human embryonic kidney HEK293T cells in which five replicates were treated with 100 nM vinblastine for 3 h and compared with five replicates of cells treated with control medium. The correction equations brought the replicates to the expected 1:1 ratios and revealed significant decreases in levels of 21 peptides upon vinblastine treatment. These equations enable accurate quantitation of small changes in peptide levels using the reductive methylation labeling approach. (AU)

• Processo FAPESP: 12/19321-9 - Caracterização proteômica e peptidômica dos venenos de aranhas brasileiras por espectrometria de massas
• Beneficiário: Alexandre Keiji Tashima
• Modalidade de apoio: Auxílio à Pesquisa - Regular