Double acylation for identification of amino-terminal peptides of proteins isolated by polyacrylamide gel electrophoresis.
We report here a method for the identification of free or blocked N-terminal peptide of in-gel digested isolated proteins. The primary amino groups of the gel-entrapped protein are blocked with normal acetic or succinic anhydride, and the protein is digested with a high-specificity protease. The generated peptides are treated with an equimolar mixture of normal and deuterated acetic anhydride. Upon mass spectrometric analysis internal peptides display a complex isotopic ion distribution while the N-terminal peptide shows a normal isotopic ion distribution. The procedure was applied to the identification of the N-terminus of individual and protein mixtures isolated by sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE).
Sanchez
Aniel
A
Department of Proteomics, Center for Genetic Engineering and Biotechnology, Havana, Cuba.
Ramos
Yassel
Y
Solano
Yanni
Y
Gonzalez
Luis Javier
LJ
Besada
Vladimir
V
Betancourt
Lazaro
L
Gil
Jeovanis
J
Alvarez
Felix
F
Rodriguez
Meilyn
M
Perez
Lincidio
L
Pujol
Merardo
M
Padron
Gabriel
G
D000215:Acylation; D000596:Amino Acids / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods; D010449:Peptide Mapping / Q000379:methods*; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods
D000596:Amino Acids; D010455:Peptides; D011506:Proteins