Research

We are developing biophysical methods using mass spectrometry to understand protein interactions. One method is PLIMSTEX (Protein Ligand Interaction by Mass Spectrometry, Titration and H/D Exchange). The figure shows the titration of fatty acid binding protein (structure on left) with the fatty-acid ligand, potassium oleate, to give a complex (on right) that now contains the fatty-acid anion. The y-axis data are mass spectrometric molecular weight measurements, and the solid line is computed from our model that gives the equilibrium constant and protection factor for the protein-ligand interaction.

A more specific approach for footprinting is to map proteins with reactions of OH radicals. In FPOP (fast photochemical oxidation of proteins), we “paint” certain surface-accessible amino-acid residues by reacting the protein in a flow tube (see figure below) with radicals generated from H₂O₂ by a pulsed KrF laser (248 nm). With appropriate scavengers, the reaction time is remarkable short (~ 1 microsec). Certain amino acids react with the radicals to become oxidized (depicted as turning from “green” to “yellow”--see figure below). PLIMSTEX and FPOP may be useful for determining protein-protein interfaces, affinities for tiny amounts of proteins in biophysics and drug discovery.

Another goal is development of Fourier transform MS. Of interest are accurate mass measurement and new trap designs (see figure below) designed to incorporate electric field compensation. Two instruments equipped with 7-tesla magnets, and soon a 12-tesla instrument for top-down proteomics are available for fundamental and applied research.

The laboratory is an NIH National Research Resource. Under this aegis, coworkers pursue method/instrument development and/or collaborate, for example, on DNA photodamage research (J Taylor), photosynthetic bacteria (R Blankenship), bacterial cell-wall analysis (J Schaefer) and antigenic peptides in immunology (E Unanue).

Selected Publications

• J.B. Sperry, X. Shi, D.L. Rempel, Y. Nishimura, S. Akashi, and M.L. Gross, “A Mass Spectrometric Approach to the Study of DNA-Binding Proteins: Interaction of Human TRF2 with Telomeric DNA,” Biochemistry, 47, 1797 (2008).
• M.R. Beck, G.T. DeKoster, D.M. Hambly, M.L. Gross, D.P. Cistola, and W.E. Goldman, W.E., “Structural features responsible for the biological stability of Histoplasma's virulence factor CBP,” Biochemistry, 47, 4427 (2008).
• A. Suri, J.J. Walters, H.W. Rohrs, M.L. Gross, and E.R. Unanue, "First Signature of Islet β-Cell-Derived Naturally Processed Peptides Selected by Diabetogenic Class II MHC Molecules," J. Immunology, 180, 3849 (2008).
• J.M. Wilcox, D.L. Rempel, and M.L. Gross, " Method of Measuring Oligonucleotide-Metal Affinities: Interactions of the Thrombin Binding Aptamer with K+ and Sr2+," Anal. Chem., 80, 2365 (2008).
• Q. Zhang and M.L. Gross, "Efficient synthesis, liquid chromatography purification, and tandem mass spectrometric characterization of estrogen-modified DNA bases," Chem. Res. Tox., 21, 1244 (2008).
• Q. Zhang, R.L. Aft and M.L. Gross, "Estrogen Carcinogenesis: Specific Identification of Estrogen-Modified Nucleobase in Breast Tissue from Women," Chem. Res. Tox., 21, 1509 (2008).
• Tingting Tu, A.D. Sauter, A.D. Sauter III, and M.L. Gross, "Improving the Signal Intensity and Sensitivity of MALDI Mass Spectrometry by Using Nanoliter Spots Deposited by Induction-Based Fluidics," J. Am. Soc. Mass Spectrom., 19, 1086 (2008).
• C.J. Hogan Jr., J.A. Carroll, H.W. Rohrs, P. Biswas, and M.L. Gross, "Charge carrier field emission determines the number of charges on native state proteins in electrospray ionization," J. Am. Chem. Soc., 130, 6926 (2008).
• D.G. Su, J.L.-F Kao, M.L. Gross, and J.-S.A. Taylor, " Structure Determination of an Interstrand-Type Cis-Anti Cyclobutane Thymine Dimer Produced in High Yield by UVB Light in an Oligodeoxynucleotide at Acidic pH," J. Am. Chem. Soc., 130, 11328 (2008).
• J.B. Sperry, J.M. Wilcox, and M.L. Gross, "Strong Anion Exchange for Studying Protein-DNA Interactions by H/D Exchange Mass Spectrometry," J. Am. Soc. Mass Spectrom., 19, 887 (2008).
• A.M. Brustkern, D.L. Rempel, and M.L. Gross, "An Electrically Compensated Trap Designed to Eighth Order for FT-ICR Mass Spectrometry," J. Am. Soc. Mass Spectrom., 19, 1281 (2008).
• G.J. Patti, J. Chen, J. Schaefer, and M.L. Gross, "Characterization of Structural Variations in the Peptidoglycan of Vancomycin-Susceptible Enterococcus faecium: Understanding Glycopeptide-Antibiotic Binding Sites Using Mass Spectrometry," J. Am. Soc. Mass Spectrom., 19, 1467 (2008).
• M.L. Gross,M.M. Zhu, and D.M. Hambly, "Two approaches to mass spectrometric protein footprinting: PLIMSTEX and FPOP," Mass Spectrometry Analysis for Protein-Protein Interactions and Dynamics (M. Chance, ed.), 243-270 (2008).
• D.M. Hambly and M.L. Gross, "Microsecond time-scale hydroxyl radical profiling of solvent-accessible protein residues," Comprehensive Anal. Chem. (J. Whitelegge, ed.), 52, 151-177 (2009).
• Q. Zhang, T. Tu, D.A. d'Avignon, and M.L. Gross, "Balance of Beneficial and Deleterious Health Effects of Quinones: A Case Study of the Chemical Properties of Genistein and Estrone Quinones," J. Am. Chem. Soc., 131, 1067 (2009).
• C.J. Hogan, J.A. Carroll, H.W. Rohrs, P. Biswas, and M.L. Gross, "Combined Charged Residue-Field Emission Model of Macromolecular Electrospray Ionization," Anal. Chem., 81, 369 (2009).