N-terminal isotope tagging with propionic anhydride: proteomic analysis of myogenic differentiation of C2C12 cells.
N-Terminal isotope tagging (NIT) is an important proteomic tool for quantifying proteins in complex mixtures. Here we describe a modified version of the isotope-coded propionylation procedure of Zhang et al. [Zhang et al., Rapid Commun. Mass Spectom. 16 (2002) 2325], which uses 'light' D0 and 'heavy' D10-propionic anhydride. The method has been extensively modified to improve both the kinetics and overall yield of propionylation. Using albumin as a model protein, the overall variation in quantification yields, calculated using several tryptic peptides, was within +/-10% (S.D. +/-0.2) error. The efficacy of the method is demonstrated by the quantitative differences obtained for vimentin in cell lysates of C2C12 myoblasts upon their myogensis to myotubules.
Nam
Hyung Wook
HW
Department of Biochemistry, College of Science, Protein Network Research Center, Yonsei University, 134 Sinchon-dong, Seoul 120-749, Republic of Korea.
Simpson
Richard
R
Kim
Yu Sam
YS
D000595:Amino Acid Sequence; D000812:Anhydrides / Q000737:chemistry*; D000818:Animals; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D003903:Deuterium; D007553:Isotope Labeling / Q000379:methods*; D051379:Mice; D008969:Molecular Sequence Data; D032446:Myoblasts / Q000166:cytology*; D010446:Peptide Fragments / Q000737:chemistry; D011422:Propionates / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D012710:Serum Albumin, Bovine; D021241:Spectrometry, Mass, Electrospray Ionization; D014357:Trypsin; D014746:Vimentin / Q000378:metabolism
D000812:Anhydrides; D010446:Peptide Fragments; D011422:Propionates; D014746:Vimentin; D012710:Serum Albumin, Bovine; D003903:Deuterium; C096126:propionic anhydride; D014357:Trypsin