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Structural Information from CW-EPR Spectra of Dipolar Coupled Nitroxide Spin Labels

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Distance Measurements in Biological Systems by EPR

Part of the book series: Biological Magnetic Resonance ((BIMR,volume 19))

Abstract

Site directed spin labeling has emerged as a powerful technique for determining structural features of proteins. By incorporating two spin labeled side chains, distances between elements of secondary structure can be determined by quantitation of spin-spin interactions between the probes. Recent advances in methods for extracting both the distance between spin labeled probes and their relative orientations are providing the capability for testing and refining accurate structural models for a wide variety of proteins. These methods take advantage of multifrequency EPR and global non-linear data analysis tools. The capabilities of current analytical methods are described for three relevant models of spin-spin interactions and directions for further refinement of these methods for future applications are described. Reliable structural information can be obtained for interelectron distances ranging from 5 to 20 Å and up to 25 Å using perdeuterated probes.

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References

  • Altenbach, C., T. Marti, H. G. Khorana, and W. L. Hubbell, 1990, Transmembrane protein structure: spin labeling of bacteriorhodopsin mutants. Science 248: 1088.

    Article  PubMed  CAS  Google Scholar 

  • Anderson, D. J., P. Hanson, J. McNulty, G. Millhauser, V. Monaco, F. Formaggio, M. Crisma, and C. Toniolo, 1999, Solution structures of TOAC-labeled trichogin GA IV peptides from allowed (g approximate to 2) and half-field electron spin resonance. J. Am. Chem. Soc. 121: 6919.

    Article  CAS  Google Scholar 

  • Barnes, J. P., Z. Liang, H. S. Mchaourab, J. H. Freed, and W. L. Hubbell, 1999, A multifrequency electron spin resonance study of T4 lysozyme dynamics. Biophys. J. 76: 3298.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Beechem, J. M., 1992, Global analysis of biochemical and biophysical data. Methods Enzymol. 210: 37.

    Article  PubMed  CAS  Google Scholar 

  • Beechem, J. M., E. Grafton, M. Ameloot, J. R. Knutson, and L. Brand, 1991, The global analysis of fluorescence intensity and anisotropy decay data: second generation theory and programs. in Topics in Fluorescence Spectroscopy ( Lakowicz, J. R., Ed.), Plenum Press, New York.

    Google Scholar 

  • Berengian, A. R., M. P. Bova, and H. S. Mchaourab, 1997, Structure and function of the conserved domain in aA-crystallin. Site-directed spin labeling identifies a ft-strand located near a subunit interface. Biochemistry 36: 9951.

    Article  PubMed  CAS  Google Scholar 

  • Berliner, L. J., J. Grunwald, H. O. Hankovsky, and K. Hideg, 1982, A novel reversible thiol- spcific spin label: papain active site labeling and inhibition. Anal. Biochem. 119: 450.

    Article  PubMed  CAS  Google Scholar 

  • Beth, A. H., B. H. Robinson, C. E. Cobb, L. R. Dalton, W. E. Trommer, J. J. Birktoft, and J. H. Park, 1984, Interactions and spatial arrangement of spin-labeled NAD+ bound to glyceraldehyde-3-phosphate dehydrogenase. J. Biol. Chem. 259: 9717.

    PubMed  CAS  Google Scholar 

  • Calvo, R., W. Hofbauer, F. Lendzian, W. Lubitz, M. L. Paddock, E. C. Abresch, R. A. Isaacson, M. Y. Ohmura, and G. Feher, 1999, EPR study of the molecular and electronic structure of the QA− QB− state of the photosynthetic reactions center in Rb. Spaeroides R26 at 35 and 94 GHz. Talk at the 22nd International EPR Symposium.

    Google Scholar 

  • Eaton, G. R., and S. S. Eaton, 1989, Resolved electron-electron spin-spin splittings in EPR spectra. in Biological Magnetic Resonance (Berliner, L. J., & Reuben, J., Eds. ), Plenum Press.

    Google Scholar 

  • Fajer, P., R. Bennet, C. Polnazek, E. Fajer, and D. Thomas, 1990, General method for multiparameter fitting of high-resolution EPR spectra using a simplex algorithm. J. Magn. Reson. 88: 111.

    CAS  Google Scholar 

  • Feix, J. B., and C. S. Klug, 1999, Site-Directed spin labeling of membrane proteins and peptide-membrane interactions. in Spin Labeling: The New Millenium (Berliner, L. J., Ed. ) pp 251.

    Google Scholar 

  • Fiori, W. R., K. M. Lundberg, and G. L. Millhauser, 1994, A Single Carboxy-Terminal Arginine Determines the Amino- Terminal Helix Conformation of an Alanine-Based Peptide. Nat. Struct. Biol. 1: 374.

    Article  PubMed  CAS  Google Scholar 

  • Fiori, W. R., G. V. Martinez, and G. L. Millhauser, 1993a, Length Dependence in Alanine-Based Peptides - the 3(10) Helix to Alpha-Helix Transition. Biophys. J. 64: A378.

    Google Scholar 

  • Fiori, W. R., S. M. Miick, and G. L. Millhauser, 1993b, Increasing Sequence Length Favors Alpha-Helix Over 3(10)-Helix in Alanine-Based Peptides - Evidence For a Length-Dependent Structural Transition. Biochemistry 32: 1 1957.

    Article  Google Scholar 

  • Fiori, W. R., and G. L. Millhauser, 1995, Exploring the Peptide 3(10)-Helix-ReversibleArrow-Alpha-Helix Equilibrium With Double-Label Electron-Spin-Resonance. Biopolymers 37: 243.

    Article  PubMed  CAS  Google Scholar 

  • Hanson, P., D. J. Anderson, G. Martinez, G. Millhauser, F. Formaggio, M. Crisma, C. Toniolo, and C. Vita, 1998, Electron spin resonance and structural analysis of water soluble, alanine-rich peptides incorporating TOAC. Mol. Phys. 95: 957.

    CAS  Google Scholar 

  • Hanson, P., G. Martinez, G. Millhauser, F. Formaggio, M. Crisma, C. Toniolo, and C. Vita, 1996a, Distinguishing helix conformations in alanine-rich peptides using the unnatural amino acid TOAC and electron spin resonance. J. Am. Chem. Soc. 118: 271.

    Article  CAS  Google Scholar 

  • Hanson, P., G. Millhauser, F. Formaggio, M. Crisma, and C. Toniolo, 1996b, ESR characterization of hexameric, helical peptides using double TOAC spin labeling. J. Am. Chem. Soc. 118: 7618.

    Article  CAS  Google Scholar 

  • Hideg, K., and O. H. Hankovszky, 1989, Chemistry of spin-labeled amino acids and peptides. Some new mono-and bifunctionalized nitroxide free radicals. in Biological Magnetic Resonance (Berliner, L. J., & Reuben, J., Eds.) pp 427, Plenum Press, New York.

    Google Scholar 

  • Hubbell, W. L., and C. Altenbach, 1994a, Investigation of structure and dynamics in membrane proteins using site-directed spin labeling. Curr. Opin. Struct. Biol. 4: 566.

    Article  CAS  Google Scholar 

  • Hubbell, W. L., and C. Altenbach, 1994b, Site-directed spin-labeling of membrane proteins. in Membrane Protein Structure ( White, S. H., Ed.) pp 224, Oxford, New York.

    Chapter  Google Scholar 

  • Hubbell, W. L., A. Gross, R. Langen, and M. A. Lietzow, 1998, Recent advances in sitedirected spin labeling of proteins. Curr. Opin. Struct. Biol. 8: 649.

    Article  PubMed  CAS  Google Scholar 

  • Hubbell, W. L., H. S. Mchaourab, C. Altenbach, and M. A. Lietzow, 1996, Watching proteins move using site-directed spin labeling. Structure 4: 779.

    Article  PubMed  CAS  Google Scholar 

  • Hustedt, E. J., and A. H. Beth, 1995, Analysis of saturation transfer electron paramagnetic resonance spectra of a spin-labeled integral membrane protein, band 3, in terms of the uniaxial rotational diffusion model. Biophys. J. 69: 1409.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Hustedt, E. J., and A. H. Beth, 1996, Determination of the orientation of a band 3 affinity spin-label relative to the membrane normal axis of the human erythrocyte. Biochemistry 35: 6944.

    Article  PubMed  CAS  Google Scholar 

  • Hustedt, E. J., and A. H. Beth, 1999, Nitroxide spin-spin interactions: Applications to protein structure and dynamics. Annu. Rev. Biophys. Biomol. Struct. 28: 129.

    Article  PubMed  CAS  Google Scholar 

  • Hustedt, E. J., C. E. Cobb, A. H. Beth, and J. M. Beechem, 1993, Measurement of rotational dynamics by the simultaneous nonlinear analysis of optical and EPR data. Biophys. J. 64: 614.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Hustedt, E. J., J. J. Kirchner, A. Spaltenstein, P. B. Hopkins, and B. H. Robinson, 1995, Monitoring DNA dynamics using spin-labels with different independent mobilities. Biochemistry 34: 4369.

    Article  PubMed  CAS  Google Scholar 

  • Hustedt, E. J., A. I. Smimov, C. F. Laub, and A. H. Beth, 1997, Molecular distances from dipolar coupled spin-labels: the global analysis of multifrequency continuous wave electron paramagnetic resonance data. Biophys. J. 74: 1861.

    Article  Google Scholar 

  • Lajzerowicz-Bonneteau, J., 1976, Molecular structure of nitroxides. in Spin Labeling: Theory and Applications ( Berliner, L. J., Ed.) pp 239, Academic Press, New York.

    Chapter  Google Scholar 

  • Martinez, G. V., and G. L. Millhauser, 1993, Alpha-Helices and 3(10)-Helices in Aqueous-Solution May Readily Be Distinguished Using FTIR Spectroscopy. Biophys. J. 64: A378.

    Google Scholar 

  • McCalley, R. C., E. J. Shimshick, and H. M. McConnell, 1972, The effect of slow rotational motion on paramagnetic resonance spectra. Chem. Phys. Lett. 13: 115.

    Article  CAS  Google Scholar 

  • Mchaourab, H. S., A. R. Berengian, and H. A. Koteiche, 1997a, Site-directed spin-labeling study of the structure and subunit interaction along a conserved sequence in the acrystallin domain of heat shock protein 27. Evidence of a conserved subunit interface. Biochemistry 36: 14627.

    Article  PubMed  CAS  Google Scholar 

  • Mchaourab, H. S., T. K#x00F3;lai, K. Hideg, and W. L. Hubbell, 1999, Motion of spin-labeled side chains in T4 lysozyme: Effects of side chain structure. Biochemistry 38: 2947.

    Article  PubMed  CAS  Google Scholar 

  • Mchaourab, H. S., M. A. Lietzow, K. Hideg, and W. L. Hubbell, 1996, Motion of spin-labeled side chains in T4 lysozyme. Correlation with protein structure and dynamics. Biochemistry 35: 7692.

    Article  PubMed  CAS  Google Scholar 

  • Mchaourab, H. S., K. J. Oh, C. J. Fang, and W. L. Hubbell, 1997b, Conformation of T4 lysozyme in solution. Hinge-bending motion and the substrate -induced conformational transition studied by site-directed spin labeling. Biochemistry 36: 307.

    Article  PubMed  CAS  Google Scholar 

  • Millhauser, G. L., 1992, Selective placement of electron resonance spin labels: new structural probes for peptrides and proteins. Trends Biochem. Sci. 17: 448.

    Article  PubMed  CAS  Google Scholar 

  • Millhauser, G. L., W. R. Fiori, and S. M. Miick, 1995, Electron spin labels. Methods Enzymol. 246: 589.

    Article  PubMed  CAS  Google Scholar 

  • Monaco, V., F. Formaggio, M. Crisma, C. Toniolo, P. Hanson, G. Millhauser, C. George, J. R. Deschamps, and J. L. Flippen-Anderson, 1999a, Determining the occurrence of a 3(10)-helix and an alpha-helix in two different segments of a lipopeptaibol antibiotic using TOAC, a nitroxide spin-labeled C-alpha-tetrasubstituted alpha-amino acid. Bioorg. Med. Chem. 7: 119.

    Article  PubMed  CAS  Google Scholar 

  • Monaco, V., F. Formaggio, M. Crisma, C. Toniolo, P. Hanson, and G. L. Millhauser, 1999b, Orientation and immersion depth of a helical lipopeptaibol in membranes using TOAC as an ESR probe. Biopolymers 50: 239.

    Article  PubMed  CAS  Google Scholar 

  • Mustafi, D., H. Joela, and M. W. Makinen, 1991, The effective position of the electronic point dipole of the nitroxyl group of spin labels determined by ENDOR spectroscopy. J. Magn. Reson. 91: 497.

    CAS  Google Scholar 

  • Mustafi, D., A. Sosa-Peinado, V. Gupta, D. Gordon, and M. W. Makinen, 2000, Endor structure of spin-labeled methylmethanethiol-sulfonate in solution and bound to TEM-1ß-lactamase. Biophys. J. 78: 382A.

    Google Scholar 

  • Park, J. H., and W. Trommer, 1989, Advantages of 15N and deuterium spin probes for biomedical electron paramagnetic resonance investigations. in Biological Magnetic Resonance (Berliner, L. J., & Reuben, J., Eds.) pp 547, Plenum Press, New York.

    Google Scholar 

  • Perozo, E., D. M. Cortes, and L. G. Cuello, 1998, Three-dimensional architecture of a K+ channel: implications for the mechanism of ion channel gating. Nat. Struct. Biol. 5: 459.

    Article  PubMed  CAS  Google Scholar 

  • Perozo, E., D. M. Cortes, and L. G. Cuello, 1999, Structural rearrangements underlying K+-channel activation gating. Science 285: 73.

    Article  PubMed  CAS  Google Scholar 

  • Press, W. H., B. P. Flannery, S. A. Teukolsky, and W. T. Vetterling (1992) Numerical Recipes in FORTRAN: the art of scientific computing., 2 ed., Cambridge University Press, New York

    Google Scholar 

  • Qu, K., J. L. Vaughn, A. Sienkiewicz, C. P. Scholes, and J. S. Fetrow, 1997, Kinetics and motional dynamics of spin-labeled yeast iso-l-cytochrome c: 1. Stopped-flow electron paramagnetic resonance as a probe for folding/unfolding of the C-terminal helix spin-labeled at cysteine 102. Biochemistry 36: 2884.

    Article  PubMed  CAS  Google Scholar 

  • Rabenstein, M. D., and Y.-K. Shin, 1995, Determination of the distance between two spin labels attached to a macromolecule. Proc. Natl. Acad. Sci. USA 92: 8239.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Robinson, B. H., L. J. Slutsky, and F. P. Auteri, 1992, Direct simulation of continuous wave electron paramagnetic resonance spectra from Brownian dynamics trajectories. J. Chem. Phys. 97: 2609.

    Article  Google Scholar 

  • Sale, K. L., K. A. Sharp, and P. G. Fajer, 1999, Predictions of spin label orientation in the crystal structure of proteins. Biophys. J. 76: A34.

    Google Scholar 

  • Schneider, D. J., and J. H. Freed, 1989, Calculating slow motional magnetic resonance spectra: a user’s guide. in Biological Magnetic Resonance (Berliner, L. J., & Reuben, J., Eds.) pp 1, Plenum Press, New York.

    Google Scholar 

  • Shin, Y.-K., C. Levinthal, F. Levinthal, and W. L. Hubbell, 1993, Colicin El binding to membranes: time-resolved studies of spin-labeled mutants. Science 259: 960.

    Article  PubMed  CAS  Google Scholar 

  • Sienkiewicz, A., A. M. d. C. Ferreira, B. Danner, and C. P. Scholes, 1999, Dielectric resonator-based flow and stopped-flow EPR with rapid field scanning: a methodology for increasing kinetic information. J. Magn. Reson. 136: 137.

    Article  PubMed  CAS  Google Scholar 

  • Steinhoff, H.-J., and W. L. Hubbell, 1996, Calculation of electron paramagnetic resonance spectra from Brownian dynamics trajectories: application to nitroxide side chains in proteins. Biophys. J. 71: 2201.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Steinhoff, H.-J., N. Radzwill, W. Thevis, V. Lenz, D. Brandenburg, A. Antson, G. Dodson, and A. Wollmer, 1997, Determination of interspin distances between spin labels attached to insulin: comparison of the electron paramagnetic resonance data with the X-ray structure. Biophys. J. 73: 3287.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Tjandra, N., and A. Bax, 1997, Direct measurement of distances and angles in biomolecules by NMR in a dilute liquid crystalline medium. Science 278: 1111.

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN, 37232, USA

    Eric J. Hustedt & Albert H. Beth

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  1. Eric J. Hustedt
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  2. Albert H. Beth
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Editors and Affiliations

  1. Ohio State University, Columbus, Ohio, USA

    Lawrence J. Berliner

  2. Department of Chemistry and Biochemistry, University of Denver, 80208, Denver, Colorado, USA

    Gareth R. Eaton  & Sandra S. Eaton  & 

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© 2002 Kluwer Academic / Plenum Publishers, New York

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Hustedt, E.J., Beth, A.H. (2002). Structural Information from CW-EPR Spectra of Dipolar Coupled Nitroxide Spin Labels. In: Berliner, L.J., Eaton, G.R., Eaton, S.S. (eds) Distance Measurements in Biological Systems by EPR. Biological Magnetic Resonance, vol 19. Springer, Boston, MA. https://doi.org/10.1007/0-306-47109-4_3

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  • DOI: https://doi.org/10.1007/0-306-47109-4_3

  • Publisher Name: Springer, Boston, MA

  • Print ISBN: 978-1-4757-0575-1

  • Online ISBN: 978-0-306-47109-4

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