title: Isotope-Encoded Carboxyl Group Footprinting for Mass Spectrometry-Based Protein Conformational Studies.
abstract: 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.
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, MO, 63130, USA.|Photosynthetic Antenna Research Center (PARC), Washington University in St. Louis, St. Louis, MO, 63130, USA.
authors.forename: Hao
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Photosynthetic Antenna Research Center (PARC), Washington University in St. Louis, St. Louis, MO, 63130, USA.|Department of Biology, Washington University in St. Louis, St. Louis, MO, 63130, USA.
authors.forename: Haijun
authors.initials: H
authors.lastname: Liu
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, MO, 63130, USA.|Photosynthetic Antenna Research Center (PARC), Washington University in St. Louis, St. Louis, MO, 63130, USA.|Department of Biology, Washington University in St. Louis, St. Louis, MO, 63130, USA.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Blankenship
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, MO, 63130, USA. mgross@wustl.edu.|Photosynthetic Antenna Research Center (PARC), Washington University in St. Louis, St. Louis, MO, 63130, USA. mgross@wustl.edu.
authors.forename: Michael L
authors.initials: ML
authors.lastname: Gross
mesh_terms: D001426:Bacterial Proteins / Q000032:analysis / Q000737:chemistry; D002853:Chromatography, Liquid; D005998:Glycine / Q000031:analogs & derivatives / Q000737:chemistry; D007553:Isotope Labeling / Q000379:methods*; D008958:Models, Molecular; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry*; D011487:Protein Conformation; D023302:Protein Footprinting / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D001426:Bacterial Proteins; D010446:Peptide Fragments; C109473:orange carotenoid protein, Synechocystis; C022010:glycine ethyl ester; D005998:Glycine
keywords: Cyanobacteria; Glycine ethyl ester (GEE); Isotope encoding; Mass spectrometry; Orange carotenoid protein; Protein footprinting