Open AccessIsotope-Encoded Carboxyl Group Footprinting for Mass Spectrometry-Based Protein Conformational Studies
Author(s) -
Hao Zhang,
Haijun Liu,
Robert E. Blankenship,
Michael L. Gross
Publication year - 2015
Publication title -
journal of the american society for mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.961
H-Index - 127
eISSN - 1879-1123
pISSN - 1044-0305
DOI - 10.1007/s13361-015-1260-5
Subject(s) - chemistry , footprinting , mass spectrometry , isotope , group (periodic table) , stable isotope labeling by amino acids in cell culture , proteomics , tandem mass spectrometry , isobaric labeling , chromatography , protein mass spectrometry , biochemistry , dna , organic chemistry , base sequence , physics , quantum mechanics , gene
We report an isotope-encoding method coupled with carboxyl-group footprinting to monitor protein conformational changes. The carboxyl groups of aspartic/glutamic acids and of the C-terminus of proteins can serve as reporters for protein conformational changes when labeled with glycine ethyl ester (GEE) mediated by carbodiimide. In the new development, isotope-encoded "heavy" and "light" GEE are used to label separately the two states of the orange carotenoid protein (OCP) from cyanobacteria. Two samples are mixed (1:1 ratio) and analyzed by a single LC-MS/MS experiment. The differences in labeling extent between the two states are represented by the ratio of the "heavy" and "light" peptides, providing information about protein conformational changes. Combining isotope-encoded MS quantitative analysis and carboxyl-group footprinting reduces the time of MS analysis and improves the sensitivity of GEE and other footprinting.