|Igor A. Kaltashov
Associate Professor, University of Massachusetts
Ph.D.: University of Maryland Baltimore County
Architecture and Dynamics of Transport Proteins
Mass spectrometry has recently evolved to become one of the major analytical tools to study biomolecular structure and function. "Soft" ionization techniques, such as electrospray ionization (ESI), provide a means to desorb intact biopolymers(proteins, oligonucleotides, polysaccharides, etc.) from solution to the gas phase. Under carefully controlled experimental conditions it is possible in many cases to preserve rather weak non-covalent biomolecular complexes and thus obtain information on binding properties in solution (e.g., protein quaternary structure, enzyme-substrate-cofactor complexes, etc.). On the other hand, rapid progress in designing/refining experimental techniques that are used to fragment mass-selected molecules in the gas phase (tandem mass spectrometry) allows direct determination of covalent structure of large biopolymers (e.g., protein primary structure) in a single experiment.
We are utilizing these capabilities of mass spectrometry to study biomolecular architecture and function. Our research is focused on: (a) developing mass spectrometry-based strategies to study higher order structure and folding dynamics of proteins, and (b) applying these techniques to study mechanisms of ligand binding/release mechanism of several transport proteins. We are currently using hydrogen/deuterium (H/D) exchange in solution in combination with ESI MS to probe the conformational stability and folding/unfolding dynamics of proteins in vitro under various conditions. Transiently populated intermediate states are detected and characterized by different degrees of backbone amide protection. Because of the high data acquisition rate, ESI MS offers a facile way to resolve these intermediates on a time scale from tens of msec to hours. The global information on the protein conformational stability and folding dynamics is complemented by local (residue-specific) information. For example, collision-activated dissociation (CAD) of protein ions is used to measure the deuterium content locally as a function of exchange time. This allows us to produce conformational stability maps of both apo- and holo-forms of proteins, thus providing valuable information on the shape of protein folding and binding funnels, from which details of protein function can be deduced.
We are also interested in exploring the structure of polypeptide and protein ions in the absence of solvent. Understanding the behavior of bioions in the gas phase is very vital for our research. For example, knowledge of mechanistic aspects of activation and dissociation of protein ions may allow us to "direct" fragmentation to specific sites (protein sub-domains, reduction of disulfide bonds, etc.) in a controlled fashion.
J.K. Hoerner, H. Xiao and I.A. Kaltashov. Structural and dynamic characteristics of a partially folded state of ubiquitin revealed by hydrogen exchange mass spectrometry . Biochemistry , 2005 , 44 , 11286 -11294
I.A. Kaltashov and A. Mohimen. Estimates of protein surface areas in solution by electrospray ionization mass spectrometry . Anal. Chem ., 2005 , 77 , 5370-5379
I.A. Kaltashov, S.J. Eyles. Mass Spectrometry in Biophysics: Conformation and Dynamics of Biomolecules . New York: John Wiley & Sons, Inc., 2005
H. Xiao, I.A. Kaltashov. Transient structural disorder as a facilitator of protein-ligand binding: native H/D exchange - mass spectrometry study of cellular retinoic acid binding protein I . J. Am. Soc. Mass Spectrom ., 2005 , 16 , 869–879
H. Xiao, J.K. Hoerner, S.J. Eyles, A. Dobo, E. Voigtman, A.I. Mel'?uk, I.A. Kaltashov. Mapping protein energy landscapes with amide hydrogen exchange and mass spectrometry. I. A generalized model for a two-state protein and comparison with experiment . Protein Sci ., 2005 , 14 , 543-557
I.A. Kaltashov. Probing protein dynamics and function under native and mildly denaturing conditions with hydrogen exchange and mass spectrometry . Int. J. Mass Spectrom ., 2005 , 240 , 249-259
I.A. Kaltashov, S.J. Eyles, H. Xiao. Combination of protein hydrogen exchange and tandem mass spectrometry as an emerging tool to probe protein structure, dynamics and function . In: Focus on Protein Research . J.W. Robinson, ed. Hauppauge, New York: Nova Science Publishers, Inc., 2004 , pp. 191-218
M. Zhang, D.R. Gumerov, A.B. Mason, I.A. Kaltashov. Indirect detection of protein-metal binding: Interaction of serum transferrin with In3+ and Bi3+ . J. Am. Soc. Mass Spectrom ., 2004 , 15 , 1658-1664
J.K. Hoerner, H. Xiao, A. Dobo, I.A. Kaltashov. Is there hydrogen scrambling in the gas phase? Energetic and structural determinants of proton mobility within protein ions . J. Am. Chem. Soc ., 2004 , 126 , 7709-7717
W.P. Griffith, I.A. Kaltashov. Highly asymmetric interactions between globin chains during hemoglobin assembly revealed by electrospray ionization mass spectrometry. Biochemistry , 2003 , 42 , 10024-10033
A. Mohimen, A. Dobo, J.K. Hoerner, I.A. Kaltashov. A chemometric approach to detection and characterization of multiple protein conformers in solution using electrospray ionization mass spectrometry . Anal. Chem ., 2003 , 75 , 4139-4147
H. Xiao, S.J. Eyles, I.A. Kaltashov. Indirect assessment of small hydrophobic ligand binding to a model protein using a combination of ESI MS and HDX/ESI MS . J. Am. Soc. Mass Spectrom ., 2003 , 14 , 506-515
I.A. Kaltashov, S.J. Eyles. Studies of biomolecular conformations and conformational dynamics with mass spectrometry. Mass Spectrom. Rev ., 2002 , 21 , 37-71
A. Dobo, I.A. Kaltashov. Detection of multiple protein conformational ensembles in solution via deconvolution of charge state distributions in ESI MS . Anal. Chem ., 2001 , 73 , 4763-4773
D. Gumerov, I.A. Kaltashov. Dynamics of iron release from transferrin N-lobe studied by electrospray ionization mass spectrometry. Anal. Chem ., 2001 , 73 , 2565-2570
S.J. Eyles, I.A. Kaltashov. Dynamics of transport proteins studied with ESI-FT ICR MS. Fourier Transform Mass Spectrometry: Principles and Applications. C.J. Cassady, G. Kruppa, eds. Marcel Dekker, submitted.
I.A. Kaltashov, A. Li, Z. Szilagyi, K. Vekey, C. Fenselau. Secondary structure of peptide ions in the gas phase evaluated by MIKE spectrometry: relevance to native conformations. Methods in Molecular Biology, Vol. 146: Protein and Peptide Analysis. New Mass Spectrometric Applications. J.R. Chapman, Ed. Humana Press, Totowa, NJ, 2000, Vol. 145, pp.133-146.
S.J. Eyles, P. Speir, G. Kruppa, L.M. Gierasch, I.A. Kaltashov. Protein conformational stability and folding/unfolding dynamics probed by Fourier transform ion cyclotron resonance mass spectrometry. J. Am. Chem. Soc., 2000, 122, 495.
S.J. Eyles, T. Dresch, L.M. Gierasch, I.A. Kaltashov. Unfolding dynamics of a b -sheet protein studied by mass spectrometry. J. Mass Spectrom., 1999, 34, 1289.
Li, C. Fenselau, I.A. Kaltashov. Stability of secondary structural elements in a solvent-free environment. II. The b -pleated sheets. Proteins: Struct. Funct. Genet. 1998, S2, 22.
I.A. Kaltashov, V.M. Doroshenko, R.J. Cotter. Gas phase H/D exchange reactions of peptide ions studied in a quadrupole ion trap mass spectrometer. Proteins: Struct. Funct. Genet. 1997, 28, 53.
I.A. Kaltashov, V. Doroshenko, R.J. Cotter, K. Takayama, N. Qureshi. Confirmation of the structure of lipid A derived from the lipopolysaccharide of Rhodobacter sphaeroides by a combination of MALDI, LSIMS, and tandem mass spectrometry. Anal. Chem. 1997, 69, 2317.
I.A. Kaltashov, R.J. Cotter, W.H. Feinstone, G.W. Ketner, A.S. Woods. Ferrichrome: surprising stability of a cyclic peptide - FeIII complex revealed by mass spectrometry. J. Am. Soc. Mass Spectrom. 1997, 8, 1070.