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Application of SAXS for the Structural Characterization of IDPs

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Book cover Intrinsically Disordered Proteins Studied by NMR Spectroscopy

Part of the book series: Advances in Experimental Medicine and Biology ((AEMB,volume 870))

Abstract

Small-angle X-ray scattering (SAXS) is a powerful structural method allowing one to study the structure, folding state and flexibility of native particles and complexes in solution and to rapidly analyze structural changes in response to variations in external conditions. New high brilliance sources and novel data analysis methods significantly enhanced resolution and reliability of structural models provided by the technique. Automation of the SAXS experiment, data processing and interpretation make solution SAXS a streamline tool for large scale structural studies in molecular biology. The method provides low resolution macromolecular shapes ab initio and is readily combined with other structural and biochemical techniques in integrative studies. Very importantly, SAXS is sensitive to macromolecular flexibility being one of the few structural techniques applicable to flexible systems and intrinsically disordered proteins (IDPs). A major recent development is the use of SAXS to study particle dynamics in solution by ensemble approaches, which allow one to quantitatively characterize flexible systems. Of special interest is the joint use of SAXS with solution NMR, given that both methods yield highly complementary structural information, in particular, for IDPs. In this chapter, we present the basics of SAXS and also consider protocols of the experiment and data analysis for different scenarios depending on the type of the studied object. These include ab initio shape reconstruction, validation of available high resolution structures and rigid body modelling for folded macromolecules and also characterisation of flexible proteins with the ensemble methods. The methods are illustrated by examples of recent applications and further perspectives of the integrative use of SAXS with NMR in the studies of IDPs are discussed.

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References

  • Bernadό P, Blackledge M (2009) A self-consistent description of the conformational behavior of chemically denatured proteins from NMR and small angle scattering. Biophys J 97(10):2839–2845. doi:10.1016/j.bpj.2009.08.044

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bernadό P, Svergun DI (2012) Structural analysis of intrinsically disordered proteins by small-angle X-ray scattering. Mol Biosyst 8(1):151–167. doi:10.1039/c1mb05275f

    Article  CAS  PubMed  Google Scholar 

  • Bernadό P, Blanchard L, Timmins P et al (2005) A structural model for unfolded proteins from residual dipolar couplings and small-angle x-ray scattering. Proc Natl Acad Sci U S A 102(47):17002–17007

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bernadό P, Mylonas E, Petoukhov MV et al (2007) Structural characterization of flexible proteins using small-angle X-ray scattering. J Am Chem Soc 129(17):5656–5664. doi:10.1021/ja069124n

    Article  CAS  PubMed  Google Scholar 

  • Bernadό P, Modig K, Grela P et al (2010) Structure and dynamics of ribosomal protein L12: An ensemble model based on SAXS and NMR relaxation. Biophys J 98(10):2374–2382. doi:S0006-3495(10)00263-8 (10.1016/j.bpj.2010.02.012)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bertini I, Giachetti A, Luchinat C et al (2010) Conformational space of flexible biological macromolecules from average data. J Am Chem Soc 132(38):13553–13558. doi:10.1021/ja1063923

    Article  CAS  PubMed  Google Scholar 

  • Blanchet CE, Svergun DI (2013) Small-angle X-ray scattering on biological macromolecules and nanocomposites in solution. Annu Rev Phys Chem 64:37–54. doi:10.1146/annurev-physchem-040412-110132

    Article  CAS  PubMed  Google Scholar 

  • Blobel J, Brath U, Bernadό P et al (2011) Protein loop compaction and the origin of the effect of arginine and glutamic acid mixtures on solubility, stability and transient oligomerization of proteins. Eur Biophys J 40(12):1327–1338. doi:10.1007/s00249-011-0686-3

    Article  CAS  PubMed  Google Scholar 

  • Boze H, Marlin T, Durand D et al (2010) Proline-rich salivary proteins have extended conformations. Biophys J 99(2):656–665. doi:10.1016/j.bpj.2010.04.050

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Feldman HJ, Hogue CW (2000) A fast method to sample real protein conformational space. Proteins 39(2):112–131

    Article  CAS  PubMed  Google Scholar 

  • Flory PJ (1953) Principles of polymer chemistry. Cornell University Press, Ithaca, United States

    Google Scholar 

  • Francis DM, Rozycki B, Koveal D et al (2011) Structural basis of p38alpha regulation by hematopoietic tyrosine phosphatase. Nat Chem Biol 7(12):916–924. doi:10.1038/nchembio.707

    Article  CAS  PubMed  Google Scholar 

  • Franke D, Svergun DI (2009) DAMMIF, a program for rapid ab-initio shape determination in small-angle scattering. J Appl Cryst 42:342–346. doi:10.1107/S0021889809000338

    Article  CAS  Google Scholar 

  • Franke D, Kikhney AG, Svergun DI (2012) Automated acquisition and analysis of small angle X-ray scattering data. Nucl Instrum Methods Phys Res 689:52–59

    Article  CAS  Google Scholar 

  • Graewert MA, Svergun DI (2013) Impact and progress in small and wide angle X-ray scattering (SAXS and WAXS). Curr Opin Struct Biol 23(5):748–754. doi:10.1016/j.sbi.2013.06.007

    Article  CAS  PubMed  Google Scholar 

  • Guinier A (1939) La diffraction des rayons X aux tres petits angles; application a l’etude de phenomenes ultramicroscopiques. Ann Phys 12:161–237

    CAS  Google Scholar 

  • Jensen MR, Markwick PRL, Meier S et al (2009) Quantitative determination of the conformational properties of partially folded and intrinsically disordered proteins using NMR dipolar couplings. Structure 17(9):1169–1185. doi:10.1016/j.str.2009.08.001

    Article  CAS  PubMed  Google Scholar 

  • Kohn JE, Millett IS, Jacob J et al (2004) Random-coil behavior and the dimensions of chemically unfolded proteins. Proc Natl Acad Sci U S A 101(34):12491–12496

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Konarev PV, Volkov VV, Sokolova AV et al (2003) PRIMUS–a Windows-PC based system for small-angle scattering data analysis. J Appl Crystallogr 36:1277–1282

    Article  CAS  Google Scholar 

  • Kozin MB, Svergun DI (2001) Automated matching of high- and low-resolution structural models. J Appl Crystallogr 34:33–41

    Article  CAS  Google Scholar 

  • Kratky O (1963) X-ray small angle scattering with substances of biological interest in diluted solutions. Progress in biophysics and molecular biology 13:105–173

    Google Scholar 

  • Krzeminski M, Marsh JA, Neale C et al (2013) Characterization of disordered proteins with ENSEMBLE. Bioinformatics 29(3):398–399. doi:10.1093/bioinformatics/bts701

    Article  CAS  PubMed  Google Scholar 

  • Le Guillou JC, Zinn-Justin J (1977) Critical exponents for the n-vector model in three dimensions from field theory. Phys Rev Lett 39(2):95–98

    Article  Google Scholar 

  • Leyrat C, Jensen MR, Ribeiro EA Jr et al (2011) The N(0)-binding region of the vesicular stomatitis virus phosphoprotein is globally disordered but contains transient αhelices. Protein Sci 20(3):542–556. doi:10.1002/pro.587

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Mandelkow EM, Mandelkow E (1998) Tau in Alzheimer’s disease. Trends Cell Biol 8(11):425–427

    Article  CAS  PubMed  Google Scholar 

  • Mattinen ML, Paakkonen K, Ikonen T et al (2002) Quaternary structure built from subunits combining NMR and small-angle x-ray scattering data. Biophys J 83(2):1177–1183

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Meier S, Grzesiek S, Blackledge M (2007) Mapping the conformational landscape of urea-denatured ubiquitin using residual dipolar couplings. J Am Chem Soc 129(31):9799–9807. doi:10.1021/ja0724339

    Article  CAS  PubMed  Google Scholar 

  • Mylonas E, Hascher A, Bernadό P et al (2008) Domain conformation of tau protein studied by solution small-angle X-ray scattering. Biochemistry 47(39):10345–10353

    Article  CAS  PubMed  Google Scholar 

  • Paoletti F, Covaceuszach S, Konarev PV et al (2009) Intrinsic structural disorder of mouse proNGF. Proteins 75(4):990–1009. doi:10.1002/prot.22311

    Article  CAS  PubMed  Google Scholar 

  • Paz A, Zeev-Ben-Mordehai T, Lundqvist M et al (2008) Biophysical characterization of the unstructured cytoplasmic domain of the human neuronal adhesion protein neuroligin 3. Biophys J 95(4):1928–1944. doi:S0006-3495(08)70151-6 (10.1529/biophysj.107.126995)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Pelikan M, Hura GL, Hammel M (2009) Structure and flexibility within proteins as identified through small angle X-ray scattering. Gen Physiol Biophys 28(2):174–189

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Petoukhov MV, Svergun DI (2005) Global rigid body modeling of macromolecular complexes against small-angle scattering data. Biophys J 89(2):1237–1250. doi:10.1529/biophysj.105.064154

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Petoukhov MV, Konarev PV, Kikhney AG et al (2007) ATSAS 2.1–towards automated and web-supported small-angle scattering data analysis. J Appl Cryst 40(s1):s223–s228

    Article  CAS  Google Scholar 

  • Porod G (1982) General theory. In: Glatter O, Kratky O (eds) Small-angle X-ray scattering. Academic, London, pp 17–51

    Google Scholar 

  • Rozycki B, Kim YC, Hummer G (2011) SAXS ensemble refinement of ESCRT-III CHMP3 conformational transitions. Structure 19(1):109–116. doi:S0969-2126(10)00395-3 (10.1016/j.str.2010.10.006)

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Stumpe MC, Grubmüller H (2007) Interaction of urea with amino acids: implications for urea-induced protein denaturation. J Am Chem Soc 129(51):16126–16131

    Article  CAS  PubMed  Google Scholar 

  • Svergun DI (1992) Determination of the regularization parameter in indirect-transform methods using perceptual criteria. J Appl Crystallogr 25:495–503

    Article  Google Scholar 

  • Svergun DI (1999) Restoring low resolution structure of biological macromolecules from solution scattering using simulated annealing. Biophys J 76(6):2879–2886

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Svergun DI (2010) Small-angle X-ray and neutron scattering as a tool for structural systems biology. Biol Chem 391(7):737–743. doi:10.1515/BC.2010.093

    Article  CAS  PubMed  Google Scholar 

  • Svergun DI, Koch MHJ (2002) Advances in structure analysis using small-angle scattering in solution. Curr Opin Struct Biol 12(5):654–660

    Article  CAS  PubMed  Google Scholar 

  • Svergun DI, Barberato C, Koch MHJ (1995) CRYSOL–a program to evaluate X-ray solution scattering of biological macromolecules from atomic coordinates. J Appl Crystallogr 28:768–773

    Article  CAS  Google Scholar 

  • Svergun DI, Petoukhov MV, Koch MHJ (2001) Determination of domain structure of proteins from X-ray solution scattering. Biophys J 80(6):2946–2953. doi:10.1016/S0006-3495(01)76260-1

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Svergun DI, Koch MHJ, Timmins PA et al (2013) Small angle X-ray and neutron scattering from solutions of biological macromolecules. OUP, Oxford

    Book  Google Scholar 

  • Tompa P (2012) On the supertertiary structure of proteins. Nat Chem Biol 8(7):597–600. doi:10.1038/nchembio.1009

    Article  CAS  PubMed  Google Scholar 

  • Volkov VV, Svergun DI (2003) Uniqueness of ab initio shape determination in small angle scattering. J Appl Crystallogr 36:860–864

    Article  CAS  Google Scholar 

  • von Ossowski I, Eaton JT, Czjzek M et al (2005) Protein disorder: conformational distribution of the flexible linker in a chimeric double cellulase. Biophys J 88(4):2823–2832

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Yang SC, Blachowicz L, Makowski L et al (2010) Multidomain assembled states of Hck tyrosine kinase in solution. Proc Natl Acad Sci U S A 107(36):15757–15762. doi:10.1073/pnas.1004569107

    Article  PubMed Central  CAS  PubMed  Google Scholar 

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Authors and Affiliations

  1. Hamburg Outstation, European Molecular Biology Laboratory, c/o DESY, Notkestrasse 85, 22603, Hamburg, Germany

    Michael Kachala, Erica Valentini & Dmitri I. Svergun

  2. Department of Chemistry, Hamburg University, Martin-Luther-King Platz 6, 20146, Hamburg, Germany

    Michael Kachala & Erica Valentini

Authors
  1. Michael Kachala
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  2. Erica Valentini
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  3. Dmitri I. Svergun
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Corresponding author

Correspondence to Michael Kachala .

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Editors and Affiliations

  1. CERM and Department of Chemistry, "Ugo Schiff" University of Florence, Florence, Italy

    Isabella C. Felli

  2. CERM and Department of Chemistry, "Ugo Schiff" University of Florence, Florence, Italy

    Roberta Pierattelli

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Kachala, M., Valentini, E., Svergun, D. (2015). Application of SAXS for the Structural Characterization of IDPs. In: Felli, I., Pierattelli, R. (eds) Intrinsically Disordered Proteins Studied by NMR Spectroscopy. Advances in Experimental Medicine and Biology, vol 870. Springer, Cham. https://doi.org/10.1007/978-3-319-20164-1_8

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