title: On the utility of predictive chromatography to complement mass spectrometry based intact protein identification.
abstract: The amino acid sequence determines the individual protein three-dimensional structure and its functioning in an organism. Therefore, "reading" a protein sequence and determining its changes due to mutations or post-translational modifications is one of the objectives of proteomic experiments. The commonly utilized approach is gradient high-performance liquid chromatography (HPLC) in combination with tandem mass spectrometry. While serving as a way to simplify the protein mixture, the liquid chromatography may be an additional analytical tool providing complementary information about the protein structure. Previous attempts to develop "predictive" HPLC for large biomacromolecules were limited by empirically derived equations based purely on the adsorption mechanisms of the retention and applicable to relatively small polypeptide molecules. A mechanism of the large biomacromolecule retention in reversed-phase gradient HPLC was described recently in thermodynamics terms by the analytical model of liquid chromatography at critical conditions (BioLCCC). In this work, we applied the BioLCCC model to predict retention of the intact proteins as well as their large proteolytic peptides separated under different HPLC conditions. The specific aim of these proof-of-principle studies was to demonstrate the feasibility of using "predictive" HPLC as a complementary tool to support the analysis of identified intact proteins in top-down, middle-down, and/or targeted selected reaction monitoring (SRM)-based proteomic experiments.
authors: Institute for Energy Problems of Chemical Physics, Russian Academy of Sciences, 119991 Moscow, Russia.
authors: Marina L
authors: ML
authors: Pridatchenko
authors: Tatyana Yu
authors: TY
authors: Perlova
authors: Hisham
authors: H
authors: Ben Hamidane
authors: Anton A
authors: AA
authors: Goloborodko
authors: Irina A
authors: IA
authors: Tarasova
authors: Alexander V
authors: AV
authors: Gorshkov
authors: Victor V
authors: VV
authors: Evreinov
authors: Yury O
authors: YO
authors: Tsybin
authors: Mikhail V
authors: MV
authors: Gorshkov
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D045304:Cytochromes c / Q000032:analysis*; D004285:Dogs; D006736:Horses; D010434:Pepsin A / Q000032:analysis*; D011487:Protein Conformation; D040901:Proteomics; D013552:Swine; D053719:Tandem Mass Spectrometry; D013816:Thermodynamics
chemical_list: D045304:Cytochromes c; D010434:Pepsin A