Online Hydrophobic Interaction Chromatography-Mass Spectrometry for Top-Down Proteomics.

Recent progress in top-down proteomics has led to a demand for mass spectrometry (MS)-compatible chromatography techniques to separate intact proteins using volatile mobile phases. Conventional hydrophobic interaction chromatography (HIC) provides high-resolution separation of proteins under nondenaturing conditions but requires high concentrations of nonvolatile salts. Herein, we introduce a series of more-hydrophobic HIC materials that can retain proteins using MS-compatible concentrations of ammonium acetate. The new HIC materials appear to function as a hybrid form of conventional HIC and reverse phase chromatography. The function of the salt seems to be preserving protein structure rather than promoting retention. Online HIC-MS is feasible for both qualitative and quantitative analysis. This is demonstrated with standard proteins and a complex cell lysate. The mass spectra of proteins from the online HIC-MS exhibit low charge-state distributions, consistent with those commonly observed in native MS. Furthermore, HIC-MS can chromatographically separate proteoforms differing by minor modifications. Hence, this new HIC-MS combination is promising for top-down proteomics.

[1]  J. Jorgenson,et al.  Improved protein recovery in reversed-phase liquid chromatography by the use of ultrahigh pressures. , 2006, Analytical Chemistry.

[2]  D. Figeys,et al.  Advancements in top-down proteomics. , 2012, Analytical chemistry.

[3]  M T Hearn,et al.  Microcalorimetric studies on the interaction mechanism between proteins and hydrophobic solid surfaces in hydrophobic interaction chromatography: effects of salts, hydrophobicity of the sorbent, and structure of the protein. , 2001, Analytical chemistry.

[4]  I. Kaltashov,et al.  Characterization of small protein aggregates and oligomers using size exclusion chromatography with online detection by native electrospray ionization mass spectrometry. , 2014, Analytical chemistry.

[5]  Christodoulos A Floudas,et al.  High Throughput Characterization of Combinatorial Histone Codes* , 2009, Molecular & Cellular Proteomics.

[6]  Ying Ge,et al.  Ultrahigh pressure fast size exclusion chromatography for top‐down proteomics , 2013, Proteomics.

[7]  Ying Ge,et al.  Top-down quantitative proteomics identified phosphorylation of cardiac troponin I as a candidate biomarker for chronic heart failure. , 2011, Journal of proteome research.

[8]  N. Kelleher,et al.  Decoding protein modifications using top-down mass spectrometry , 2007, Nature Methods.

[9]  J. C. Tran,et al.  Intact proteome fractionation strategies compatible with mass spectrometry , 2011, Expert review of proteomics.

[10]  Richard D. LeDuc,et al.  Deep and quantitative top-down proteomics in clinical and translational research , 2014, Expert review of proteomics.

[11]  M. Schmuck,et al.  Optimization of preparative hydrophobic interaction chromatographic purification methods. , 1986, Journal of chromatography.

[12]  Jiang Zhang,et al.  Integrating Native Mass Spectrometry and Top-Down MS for Defining Protein Interactions Important in Biology and Medicine. , 2013, Mass spectrometry.

[13]  Richard D. LeDuc,et al.  Mapping Intact Protein Isoforms in Discovery Mode Using Top Down Proteomics , 2011, Nature.

[14]  T. Tetaz,et al.  Hydrophilic interaction chromatography of intact, soluble proteins. , 2011, Journal of chromatography. A.

[15]  M. Nilges,et al.  Posttranslational Modification of Pili upon Cell Contact Triggers N. meningitidis Dissemination , 2011, Science.

[16]  Charles Ansong,et al.  Top-down proteomics reveals a unique protein S-thiolation switch in Salmonella Typhimurium in response to infection-like conditions , 2013, Proceedings of the National Academy of Sciences.

[17]  Weidong Cui,et al.  Top-down mass spectrometry: recent developments, applications and perspectives. , 2011, The Analyst.

[18]  David L Tabb,et al.  Characterization of the 70S Ribosome from Rhodopseudomonas palustris using an integrated "top-down" and "bottom-up" mass spectrometric approach. , 2004, Journal of proteome research.

[19]  J. O’Connell,et al.  Protein unfolding during reversed-phase chromatography: II. Role of salt type and ionic strength. , 2001, Biotechnology and bioengineering.

[20]  A. Jungbauer,et al.  Hydrophobic interaction chromatography of proteins. III. Unfolding of proteins upon adsorption. , 2005, Journal of chromatography. A.

[21]  Joe Carroll,et al.  Definition of the mitochondrial proteome by measurement of molecular masses of membrane proteins , 2006, Proceedings of the National Academy of Sciences.

[22]  Ying Ge,et al.  Top-down high-resolution mass spectrometry of cardiac myosin binding protein C revealed that truncation alters protein phosphorylation state , 2009, Proceedings of the National Academy of Sciences.

[23]  A. Alpert,et al.  Effective Protein Separation by Coupling Hydrophobic Interaction and Reverse Phase Chromatography for Top-down Proteomics , 2014, Analytical chemistry.

[24]  B. Chait.  Mass Spectrometry: Bottom-Up or Top-Down? , 2006, Science.

[25]  W. Y. Chen,et al.  Microcalorimetric studies of interactions between proteins and hydrophobic ligands in hydrophobic interaction chromatography: effects of ligand chain length, density and the amount of bound protein. , 2000, Journal of chromatography. A.

[26]  Ronald J Moore,et al.  Enhanced top-down characterization of histone post-translational modifications , 2012, Genome Biology.

[27]  A. Alpert.  High-performance hydrophobic-interaction chromatography of proteins on a series of poly(alkyl aspart-amide)-silicas , 1986 .

[28]  Igor A Kaltashov,et al.  Characterization of intact protein conjugates and biopharmaceuticals using ion-exchange chromatography with online detection by native electrospray ionization mass spectrometry and top-down tandem mass spectrometry. , 2015, Analytical chemistry.

[29]  Tawnya G. Flick,et al.  Solution additives that desalt protein ions in native mass spectrometry. , 2012, Analytical chemistry.

[30]  A. Cheema,et al.  Small Changes Huge Impact: The Role of Protein Posttranslational Modifications in Cellular Homeostasis and Disease , 2011, Journal of amino acids.

[31]  L. Mazzarella,et al.  Selective deamidation of ribonuclease A. Isolation and characterization of the resulting isoaspartyl and aspartyl derivatives. , 1993, The Journal of biological chemistry.

[32]  Ying Ge,et al.  Augmented Phosphorylation of Cardiac Troponin I in Hypertensive Heart Failure* , 2011, The Journal of Biological Chemistry.

[33]  Ying Ge,et al.  Top-down Proteomics Reveals Concerted Reductions in Myofilament and Z-disc Protein Phosphorylation after Acute Myocardial Infarction* , 2014, Molecular & Cellular Proteomics.

[34]  Alois Jungbauer,et al.  Hydrophobic interaction chromatography of proteins IV. Kinetics of protein spreading. , 2007, Journal of chromatography. A.

[35]  Gianluca Damonte,et al.  Achievements and perspectives of top‐down proteomics , 2010, Proteomics.

[36]  N. Kelleher,et al.  The emergence of top-down proteomics in clinical research , 2013, Genome Medicine.

[37]  J. Jorgenson,et al.  Prefractionation of intact proteins by reversed-phase and anion-exchange chromatography for the differential proteomic analysis of Saccharomyces cerevisiae. , 2013, Journal of proteome research.

[38]  A. Heck.  Native mass spectrometry: a bridge between interactomics and structural biology , 2008, Nature Methods.

[39]  Sudhir Srivastava,et al.  Posttranslational Protein Modifications , 2006, Molecular & Cellular Proteomics.

[40]  Ying Ge,et al.  Three dimensional liquid chromatography coupling ion exchange chromatography/hydrophobic interaction chromatography/reverse phase chromatography for effective protein separation in top-down proteomics. , 2015, Analytical chemistry.

[41]  J. Queiroz,et al.  Hydrophobic interaction chromatography of proteins. , 2001, Journal of biotechnology.

[42]  Mohammed Shameem,et al.  Separation of mAbs molecular variants by analytical hydrophobic interaction chromatography HPLC: overview and applications , 2014, mAbs.

[43]  N. Kelleher,et al.  Size-sorting combined with improved nanocapillary liquid chromatography-mass spectrometry for identification of intact proteins up to 80 kDa. , 2010, Analytical chemistry.

[44]  S. Way,et al.  Proteolytic elimination of N-myristoyl modifications by the Shigella virulence factor IpaJ , 2013, Nature.

[45]  Steven M. Patrie,et al.  Sensitive and reproducible intact mass analysis of complex protein mixtures with superficially porous capillary reversed-phase liquid chromatography mass spectrometry. , 2011, Analytical chemistry.

[46]  Lloyd M. Smith,et al.  Proteoform: a single term describing protein complexity , 2013, Nature Methods.

[47]  Ying Ge,et al.  Top‐down proteomics in health and disease: Challenges and opportunities , 2014, Proteomics.

[48]  B. Garcia.  What does the future hold for top down mass spectrometry? , 2010, Journal of the American Society for Mass Spectrometry.