Sample Preparation and Relative Quantitation using Reductive Methylation of Amines for Peptidomics Studies

Autor(es)

Correa, Claudia Neves ^{[1, 2]} ; Fiametti, Louise Oliveira ^[1] ; Mazzi Esquinca, Maria Eduarda ^[1] ; de Castro, Leandro Mantovani ^{[1, 2]}

Afiliação do(s) autor(es)

^[1] Sao Paulo State Univ UNESP, Biosci Inst, Dept Biol & Environm Sci, Presidente Prudente, SP - Brazil

^[2] Sao Paulo State Univ UNESP, Biodivers Coastal Environm Postgrad Program, Biosci Inst, Presidente Prudente, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: JOVE-JOURNAL OF VISUALIZED EXPERIMENTS; n. 177 NOV 2021.
• Citações Web of Science: 0

Resumo

Peptidomics can be defined as the qualitative and quantitative analysis of peptides in a biological sample. Its main applications include identifying the peptide biomarkers of disease or environmental stress, identifying neuropeptides, hormones, and bioactive intracellular peptides, discovering antimicrobial and nutraceutical peptides from protein hydrolysates, and can be used in studies to understand the proteolytic processes. The recent advance in sample preparation, separation methods, mass spectrometry techniques, and computational tools related to protein sequencing has contributed to the increase of the identified peptides number and peptidomes characterized. Peptidomic studies frequently analyze peptides that are naturally generated in cells. Here, a sample preparation protocol based on heat-inactivation is described, which eliminates protease activity, and extraction with mild conditions, so there is no peptide bonds cleavage. In addition, the relative quantitation of peptides using stable isotope labeling by reductive methylation of amines is also shown. This labeling method has some advantages as the reagents are commercially available, inexpensive compared to others, chemically stable, and allows the analysis of up to five samples in a single LC-MS run. (AU)

• Processo FAPESP: 21/01286-1 - Transcriptoma e peptidoma de tentáculos da anêmona marinha Bunodosoma caissarum
• Beneficiário: Maria Eduarda Mazzi Esquinca
• Modalidade de apoio: Bolsas no Brasil - Iniciação Científica
• Processo FAPESP: 19/17433-3 - Avaliação dos efeitos citotóxicos ou neuroprotetores de peptídeos alterados no sistema nervoso central de zebrafish (Danio rerio) na Doença de Parkinson
• Beneficiário: Louise Oliveira Fiametti
• Modalidade de apoio: Bolsas no Brasil - Iniciação Científica
• Processo FAPESP: 19/16023-6 - Revelando dados de transcriptoma e proteômica para caracterização de peptidomas e peptídeos bioativos em organismos da fauna acompanhante
• Beneficiário: Leandro Mantovani de Castro
• Modalidade de apoio: Auxílio à Pesquisa - Regular