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Crystallographic Studies of Intermediate Filament Proteins

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Fibrous Proteins: Structures and Mechanisms

Part of the book series: Subcellular Biochemistry ((SCBI,volume 82))

Abstract

Intermediate filaments (IFs), together with microtubules and actin microfilaments, are the three main cytoskeletal components in metazoan cells. IFs are formed by a distinct protein family, which is made up of 70 members in humans. Most IF proteins are tissue- or organelle-specific, which includes lamins, the IF proteins of the nucleus. The building block of IFs is an elongated dimer, which consists of a central α-helical ‘rod’ domain flanked by flexible N- and C-terminal domains. The conserved rod domain is the ‘signature feature’ of the IF family. Bioinformatics analysis reveals that the rod domain of all IF proteins contains three α-helical segments of largely conserved length, interconnected by linkers. Moreover, there is a conserved pattern of hydrophobic repeats within each segment, which includes heptads and hendecads. This defines the presence of both left-handed and almost parallel coiled-coil regions along the rod length. Using X-ray crystallography on multiple overlapping fragments of IF proteins, the atomic structure of the nearly complete rod domain has been determined. Here, we discuss some specific challenges of this procedure, such as crystallization and diffraction data phasing by molecular replacement. Further insights into the structure of the coiled coil and the terminal domains have been obtained using electron paramagnetic resonance measurements on the full-length protein, with spin labels attached at specific positions. This atomic resolution information, as well as further interesting findings, such as the variation of the coiled-coil stability along the rod length, provide clues towards interpreting the data on IF assembly, collected by a range of methods. However, a full description of this process at the molecular level is not yet at hand.

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Abbreviations

EPR:

Electron Paramagnetic Resonance

IF:

Intermediate Filament

MF:

Microfilament

MR:

Molecular Replacement

MT:

Microtubule

RMSD:

Root-Mean-Square Deviation

SDSL:

Site-Directed Spin Labelling

ULF:

Unit-Length Filament

References

  • Ausmees N, Kuhn JR, Jacobs-Wagner C (2003) The bacterial cytoskeleton: an intermediate filament-like function in cell shape. Cell 115(6):705–713

    Article  CAS  PubMed  Google Scholar 

  • Aziz A, Hess JF, Budamagunta MS, Voss JC, Fitzgerald PG (2010) Site-directed spin labeling and electron paramagnetic resonance determination of vimentin head domain structure. J Biol Chem 285(20):15278–15285

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Aziz A, Hess JF, Budamagunta MS, Voss JC, Kuzin AP, Huang YJ, Xiao R, Montelione GT, FitzGerald PG, Hunt JF (2012) The structure of vimentin linker 1 and rod 1B domains characterized by site-directed spin-labeling electron paramagnetic resonance (SDSL-EPR) and X-ray crystallography. J Biol Chem 287(34):28349–28361

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bailey TL, Williams N, Misleh C, Li WW (2006) MEME: discovering and analyzing DNA and protein sequence motifs. Nucleic Acids Res 34(suppl 2):W369–W373

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bollati M, Barbiroli A, Favalli V, Arbustini E, Charron P, Bolognesi M (2012) Structures of the lamin A/C R335W and E347K mutants: implications for dilated cardiolaminopathies. Biochem Biophys Res Commun 418(2):217–221

    Article  CAS  PubMed  Google Scholar 

  • Brown JH, Cohen C, Parry DAD (1996) Heptad breaks in alpha-helical coiled coils: stutters and stammers. Proteins 26(2):134–145

    Article  CAS  PubMed  Google Scholar 

  • Chernyatina AA, Strelkov SV (2012) Stabilization of vimentin coil2 fragment via an engineered disulfide. J Struct Biol 177(1):46–53

    Article  CAS  PubMed  Google Scholar 

  • Chernyatina AA, Nicolet S, Aebi U, Herrmann H, Strelkov SV (2012) Atomic structure of the vimentin central alpha-helical domain and its implications for intermediate filament assembly. Proc Natl Acad Sci U S A 109(34):13620–13625

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chernyatina AA, Guzenko D, Strelkov SV (2015) Intermediate filament structure: the bottom-up approach. Curr Opin Cell Biol 32C:65–72

    Article  Google Scholar 

  • Chernyatina AA, Hess JF, Guzenko D, Voss JC, Strelkov SV (2016) How to study intermediate filaments in atomic detail. Methods Enzymol 568:3–33

    Article  PubMed  Google Scholar 

  • Dhe-Paganon S, Werner ED, Chi YI, Shoelson SE (2002) Structure of the globular tail of nuclear lamin. J Biol Chem 277(20):17381–17384

    Article  CAS  PubMed  Google Scholar 

  • Dittmer TA, Misteli T (2011) The lamin protein family. Genome Biol 12(5):222

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Drozdetskiy A, Cole C, Procter J, Barton GJ (2015) JPred4: a protein secondary structure prediction server. Nucleic Acids Res 43(W1):W389–W394

    Article  PubMed  PubMed Central  Google Scholar 

  • Goldie KN, Wedig T, Mitra AK, Aebi U, Herrmann H, Hoenger A (2007) Dissecting the 3-D structure of vimentin intermediate filaments by cryo-electron tomography. J Struct Biol 158(3):378–385

    Article  CAS  PubMed  Google Scholar 

  • Goldman RD, Khuon S, Chou YH, Opal P, Steinert PM (1996) The function of intermediate filaments in cell shape and cytoskeletal integrity. J Cell Biol 134(4):971–983

    Article  CAS  PubMed  Google Scholar 

  • Grigoryan G, DeGrado WF (2011) Probing designability via a generalized model of helical bundle geometry. J Mol Biol 405(4):1079–1100

    Article  CAS  PubMed  Google Scholar 

  • Gruber M, Lupas AN (2003) Historical review: another 50th anniversary–new periodicities in coiled coils. Trends Biochem Sci 28(12):679–685

    Article  CAS  PubMed  Google Scholar 

  • Guindon S, Dufayard J-F, Lefort V, Anisimova M, Hordijk W, Gascuel O (2010) New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst Biol 59(3):307–321

    Article  CAS  PubMed  Google Scholar 

  • Herrmann H, Haner M, Brettel M, Muller SA, Goldie KN, Fedtke B, Lustig A, Franke WW, Aebi U (1996) Structure and assembly properties of the intermediate filament protein vimentin: the role of its head, rod and tail domains. J Mol Biol 264(5):933–953

    Article  CAS  PubMed  Google Scholar 

  • Herrmann H, Aebi U (2004) Intermediate filaments: molecular structure, assembly mechanism, and integration into functionally distinct intracellular Scaffolds. Annu Rev Biochem 73:749–789

    Article  CAS  PubMed  Google Scholar 

  • Herrmann H, Kreplak L, Aebi U (2004) Isolation, characterization, and in vitro assembly of intermediate filaments. Methods Cell Biol 78:3–24

    Article  CAS  PubMed  Google Scholar 

  • Herrmann H, Bar H, Kreplak L, Strelkov SV, Aebi U (2007) Intermediate filaments: from cell architecture to nanomechanics. Nat Rev Mol Cell Biol 8(7):562–573

    Article  CAS  PubMed  Google Scholar 

  • Hess JF, Budamagunta MS, Voss JC, FitzGerald PG (2004) Structural characterization of human vimentin rod 1 and the sequencing of assembly steps in intermediate filament formation in vitro using site-directed spin labeling and electron paramagnetic resonance. J Biol Chem 279(43):44841–44846

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hess JF, Budamagunta MS, Shipman RL, FitzGerald PG, Voss JC (2006) Characterization of the linker 2 region in human vimentin using site-directed spin labeling and electron paramagnetic resonance. Biochemistry 45(39):11737–11743

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hess JF, Budamagunta MS, Aziz A, FitzGerald PG, Voss JC (2013) Electron paramagnetic resonance analysis of the vimentin tail domain reveals points of order in a largely disordered region and conformational adaptation upon filament assembly. Protein Sci 22(1):47–55

    Article  CAS  PubMed  Google Scholar 

  • Ivaska J, Pallari HM, Nevo J, Eriksson JE (2007) Novel functions of vimentin in cell adhesion, migration, and signaling. Exp Cell Res 313(10):2050–2062

    Article  CAS  PubMed  Google Scholar 

  • Jones DT, Cozzetto D (2015) DISOPRED3: precise disordered region predictions with annotated protein-binding activity. Bioinformatics 31(6):857–863

    Article  CAS  PubMed  Google Scholar 

  • Kapinos LE, Burkhard P, Herrmann H, Aebi U, Strelkov SV (2011) Simultaneous formation of right- and left-handed anti-parallel coiled-coil interfaces by a coil2 fragment of human lamin A. J Mol Biol 408(1):135–146

    Article  CAS  PubMed  Google Scholar 

  • Kaufmann KW, Lemmon GH, DeLuca SL, Sheehan JH, Meiler J (2010) Practically useful: what the Rosetta protein modeling suite can do for you. Biochemistry 49(14):2987–2998

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kuhnel K, Jarchau T, Wolf E, Schlichting I, Walter U, Wittinghofer A, Strelkov SV (2004) The VASP tetramerization domain is a right-handed coiled coil based on a 15-residue repeat. Proc Natl Acad Sci U S A 101(49):17027–17032

    Article  PubMed  PubMed Central  Google Scholar 

  • Lee CH, Kim MS, Chung BM, Leahy DJ, Coulombe PA (2012) Structural basis for heteromeric assembly and perinuclear organization of keratin filaments. Nat Struct Mol Biol 19(7):707–715

    Article  CAS  PubMed  Google Scholar 

  • Lupas A, Van Dyke M, Stock J (1991) Predicting coiled coils from protein sequences. Science 252(5009):1162–1164

    Article  CAS  PubMed  Google Scholar 

  • MacPhee CE, Woolfson DN (2004) Engineered and designed peptide-based fibrous biomaterials. Curr Opinion Solid State Mater Sci 8(2):141–149

    Article  CAS  Google Scholar 

  • Meier M, Padilla GP, Herrmann H, Wedig T, Hergt M, Patel TR, Stetefeld J, Aebi U, Burkhard P (2009) Vimentin coil 1A - A molecular switch involved in the initiation of filament elongation. J Mol Biol 390(2):245–261

    Article  CAS  PubMed  Google Scholar 

  • Mencarelli C, Ciolfi S, Caroti D, Lupetti P, Dallai R (2011) Isomin: a novel cytoplasmic intermediate filament protein from an arthropod species. BMC Biol 9:17

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mucke N, Wedig T, Burer A, Marekov LN, Steinert PM, Langowski J, Aebi U, Herrmann H (2004) Molecular and biophysical characterization of assembly-starter units of human vimentin. J Mol Biol 340(1):97–114

    Article  CAS  PubMed  Google Scholar 

  • Nicolet S, Herrmann H, Aebi U, Strelkov SV (2010) Atomic structure of vimentin coil 2. J Struct Biol 170(2):369–376

    Article  CAS  PubMed  Google Scholar 

  • Parry DAD (2006) Hendecad repeat in segment 2A and linker L2 of intermediate filament chains implies the possibility of a right-handed coiled-coil structure. J Struct Biol 155(2):370–374

    Article  CAS  PubMed  Google Scholar 

  • Parry DAD, Fraser RD, Squire JM (2008) Fifty years of coiled-coils and alpha-helical bundles: a close relationship between sequence and structure. J Struct Biol 163(3):258–269

    Article  CAS  PubMed  Google Scholar 

  • Patterson AL (1935) A direct method for the determination of the components of interatomic distances in crystals. Zeitschrift für Kristallographie-Crystalline Materials 90(1):517–542

    CAS  Google Scholar 

  • Pekny M, Lane EB (2007) Intermediate filaments and stress. Exp Cell Res 313(10):2244–2254

    Article  CAS  PubMed  Google Scholar 

  • Peter A, Stick R (2015) Evolutionary aspects in intermediate filament proteins. Curr Opin Cell Biol 32:48–55

    Article  CAS  PubMed  Google Scholar 

  • Petersen B, Petersen TN, Andersen P, Nielsen M, Lundegaard C (2009) A generic method for assignment of reliability scores applied to solvent accessibility predictions. BMC Struct Biol 9:51

    Article  PubMed  PubMed Central  Google Scholar 

  • Potapov V, Kaplan JB, Keating AE (2015) Data-driven prediction and design of bZIP coiled-coil interactions. PLoS Comput Biol 11(2):e1004046

    Article  PubMed  PubMed Central  Google Scholar 

  • Rämisch S, Lizatović R, André I (2015) Automated de novo phasing and model building of coiled-coil proteins. Acta Crystallogr D Biol Crystallogr 71(3):606–614

    Article  PubMed  Google Scholar 

  • Robert A, Hookway C, Gelfand VI (2016) Intermediate filament dynamics: what we can see now and why it matters. BioEssays 38(3):232–243

    Article  PubMed  Google Scholar 

  • Rossmann MG (1990) The molecular replacement method. Acta Crystallogr A: Found Crystallogr 46(2):73–82

    Article  Google Scholar 

  • Ruan J, Xu C, Bian C, Lam R, Wang JP, Kania J, Min JR, Zang JY (2012) Crystal structures of the coil 2B fragment and the globular tail domain of human lamin B1. FEBS Lett 586(4):314–318

    Article  CAS  PubMed  Google Scholar 

  • Satelli A, Li S (2011) Vimentin in cancer and its potential as a molecular target for cancer therapy. Cell Mol Life Sci 68(18):3033–3046

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Scally SW, Petersen J, Law SC, Dudek NL, Nel HJ, Loh KL, Wijeyewickrema LC, Eckle SBG, van Heemst J, Pike RN, McCluskey J, Toes RE, La Gruta NL, Purcell AW, Reid HH, Thomas R, Rossjohn J (2013) A molecular basis for the association of the HLA-DRB1 locus, citrullination, and rheumatoid arthritis. J Exp Med 210(12):2569–2582

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Snider NT, Omary MB (2014) Post-translational modifications of intermediate filament proteins: mechanisms and functions. Nat Rev Mol Cell Biol 15(3):163–177

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Steinert PM, Roop DR (1988) Molecular and cellular biology of intermediate filaments. Annu Rev Biochem 57:593–625

    Article  CAS  PubMed  Google Scholar 

  • Steinert PM, Marekov LN, Parry DAD (1993) Diversity of intermediate filament structure. Evidence that the alignment of coiled-coil molecules in vimentin is different from that in keratin intermediate filaments. J Biol Chem 268(33):24916–24925

    CAS  PubMed  Google Scholar 

  • Steinert PM, Marekov LN, Parry DAD (1999) Molecular parameters of type IV α-internexin and type IV-type III α-internexin-vimentin copolymer intermediate filaments. J Biol Chem 274(3):1657–1666

    Article  CAS  PubMed  Google Scholar 

  • Strelkov SV, Burkhard P (2002) Analysis of alpha-helical coiled coils with the program TWISTER reveals a structural mechanism for stutter compensation. J Struct Biol 137(1–2):54–64

    Article  CAS  PubMed  Google Scholar 

  • Strelkov SV, Herrmann H, Geisler N, Lustig A, Ivaninskii S, Zimbelmann R, Burkhard P, Aebi U (2001) Divide-and-conquer crystallographic approach towards an atomic structure of intermediate filaments. J Mol Biol 306(4):773–781

    Article  CAS  PubMed  Google Scholar 

  • Strelkov SV, Herrmann H, Geisler N, Wedig T, Zimbelmann R, Aebi U, Burkhard P (2002) Conserved segments 1A and 2B of the intermediate filament dimer: their atomic structures and role in filament assembly. EMBO J 21(6):1255–1266

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Strelkov SV, Schumacher J, Burkhard P, Aebi U, Herrmann H (2004) Crystal structure of the human lamin A coil 2B dimer: implications for the head-to-tail association of nuclear lamins. J Mol Biol 343(4):1067–1080

    Article  CAS  PubMed  Google Scholar 

  • Szeverenyi I, Cassidy AJ, Chung CW, Lee BT, Common JE, Ogg SC, Chen H, Sim SY, Goh WLR, Ng KW, Simpson JA, Chee LL, Eng GH, Li B, Lunny DP, Chuon D, Venkatesh A, Khoo KH, McLean WHI, Lim YP, Lane EB (2008) The Human Intermediate Filament Database: comprehensive information on a gene family involved in many human diseases. Hum Mutat 29(3):351–360

    Article  CAS  PubMed  Google Scholar 

  • Taylor GL (2010) Introduction to phasing. Acta Crystallogr D Biol Crystallogr 66(Pt 4):325–338

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Toivola DM, Strnad P, Habtezion A, Omary MB (2010) Intermediate filaments take the heat as stress proteins. Trends Cell Biol 20(2):79–91

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vincent TL, Green PJ, Woolfson DN (2013) LOGICOIL—multi-state prediction of coiled-coil oligomeric state. Bioinformatics 29(1):69–76

    Article  CAS  PubMed  Google Scholar 

  • Wickert U, Mucke N, Wedig T, Muller SA, Aebi U, Herrmann H (2005) Characterization of the in vitro co-assembly process of the intermediate filament proteins vimentin and desmin: mixed polymers at all stages of assembly. Eur J Cell Biol 84(2–3):379–391

    Article  CAS  PubMed  Google Scholar 

  • Wood CW, Bruning M, Ibarra AÁ, Bartlett GJ, Thomson AR, Sessions RB, Brady RL, Woolfson DN (2014) CCBuilder: an interactive web-based tool for building, designing and assessing coiled-coil protein assemblies. Bioinformatics 30:3029–3035 btu502

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors are thankful to Prof. David A.D. Parry and Dr. Oleg A. Klychnikov for critical reading of the manuscript, and to Prof. Harald Herrmann for fruitful discussions.

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Authors and Affiliations

  1. Laboratory for Biocrystallography, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, Leuven, Belgium

    Dmytro Guzenko, Anastasia A. Chernyatina & Sergei V. Strelkov

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  1. Dmytro Guzenko
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  2. Anastasia A. Chernyatina
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  3. Sergei V. Strelkov
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Correspondence to Sergei V. Strelkov .

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Editors and Affiliations

  1. Institute of Fundamental Sciences and Riddet Institute, Massey University, Palmerston North, New Zealand

    David A.D. Parry

  2. Muscle Contraction Group, School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom

    John M. Squire

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Guzenko, D., Chernyatina, A.A., Strelkov, S.V. (2017). Crystallographic Studies of Intermediate Filament Proteins. In: Parry, D., Squire, J. (eds) Fibrous Proteins: Structures and Mechanisms. Subcellular Biochemistry, vol 82. Springer, Cham. https://doi.org/10.1007/978-3-319-49674-0_6

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