Nanomechanics of Proteins, Both Folded and Disordered

  • Rubén Hervás
  • Albert Galera-Prat
  • Àngel Gómez-Sicilia
  • Fernando Losada-Urzáiz
  • María del Carmen Fernández
  • Débora Fernández-Bravo
  • Elena Santana
  • Clara Barrio-García
  • Carolina Melero
  • Mariano Carrión-Vázquez
Part of the Biophysics for the Life Sciences book series (BIOPHYS, volume 2)


Single-molecule techniques have recently provided a versatile tool for imaging and manipulating protein molecules one at a time, enabling us to address important biological questions in key areas of cell biology (e.g., cell adhesion and signaling, neurodegeneration) and protein science (e.g., protein folding, protein structure and stability, catalysis, protein evolution, conformational polymorphism, and amyloidogenesis). One of these techniques, single-molecule force spectroscopy based on atomic force microscopy, combined with theoretical/computational approaches and protein engineering, has allowed unprecedented progress in characterizing and understanding at the molecular level the mechanical properties of biomolecules, particularly those of proteins, which has recently opened the new, exciting and fast-growing research field of protein nanomechanics. The aim of this review is to describe the principles of this methodology and to discuss the main achievements in this field, with special emphasis on its emerging application to the analysis of intrinsically disordered proteins.



Atomic force microscopy



Amyloid beta




Disorder in disorder




Dihydrofolate reductase


Dimethyl sulfoxide




Unfolding force


B1 immunoglobulin binding domain of streptococcal protein G


Huntington’s disease




Intrinsically disordered protein




Contour length




Maltose binding protein




Avogadro’s number


Nicotinamide adenine dihydrogen phosphate




Nuclear magnetic resonance


Persistence length




Plasmid for force spectroscopy


Protein kinase A




PolyQ binding peptide 1


Random coil


Steered molecular dynamics


Single-molecule fluorescence


Single-molecule force spectroscopy


Single-molecule techniques


Scanning probe microscopy


Disulfide bond


Scanning tunneling microscope




Third fibronectin type III domain


Seventh FnIII domain of human tenascin-X


Transition state


Worm-like chain


Alpha synuclein


Contour length increase


  1. Ainavarapu SRK, Li L, Badilla CL, Fernandez JM (2005) Ligand binding modulates the mechanical stability of dihydrofolate reductase. Biophys J 89:3337–3344PubMedGoogle Scholar
  2. Ainavarapu SRK, Brujic J, Huang HH, Wiita AP, Lu H, Li L, Walther KA, Carrión-Vázquez M, Li H, Fernandez JM (2007) Contour length and refolding rate of a small protein controlled by engineering disulfide bonds. Biophys J 92:22–233Google Scholar
  3. Ainavarapu SRK, Wiita AP, Dougan L, Uggerud E, Fernandez JM (2008a) Single-molecule force spectroscopy measurements of bond elongation during a bimolecular reaction. J Am Chem Soc 130:6479–6487Google Scholar
  4. Ainavarapu SRK, Wiita AP, Huang HH, Fernandez JM (2008b) A single-molecule assay to directly identify solvent-accessible disulfide bonds and probe their effect on protein folding. J Am Chem Soc 130:436–437PubMedGoogle Scholar
  5. Aioanei D, Lv S, Tessari I, Rampioni A, Bubacco L, Li H, Samorì B, Brucale M (2011a) Single-molecule-level evidence for the osmophobic effect. Angew Chem Int Ed 50:4394–4397Google Scholar
  6. Aioanei D, Tessari I, Bubacco L, Samorì B, Brucale M (2011b) Observing the osmophobic effect in action at the single molecule level. Proteins 79:2214–2223PubMedGoogle Scholar
  7. Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002) Molecular biology of the cell, 4th edn. Garland Science, New York, NYGoogle Scholar
  8. Alegre-Cebollada J, Kosuri P, Rivas-Pardo JA, Fernández JM (2011) Direct observation of disulfide isomerization in a single protein. Nat Chem 3:882–887PubMedGoogle Scholar
  9. Arslan PE, Mulligan VK, Ho S, Chakrabartty A (2010) Conversion of Aβ42 into a folded soluble native-like protein using a semi-random library of amphipathic helices. J Mol Biol 396:1284–1294PubMedGoogle Scholar
  10. Baldock C, Oberhauser AF, Ma L, Lammie D, Siegler V, Mithieux SM, Tu Y, Chow JY, Suleman F, Malfois M, Rogers S, Guo L, Irving TC, Wess TJ, Weiss AS (2011) Shape of tropoelastin, the highly extensible protein that controls human tissue elasticity. Proc Natl Acad Sci USA 108:4322–4327PubMedGoogle Scholar
  11. Bell GI (1978) Models for the specific adhesion of cells to cells. Science 200:618–627PubMedGoogle Scholar
  12. Bertz M, Rief M (2009) Ligand binding mechanics of maltose binding protein. J Mol Biol 393:1097–1105PubMedGoogle Scholar
  13. Best RB, Hummer G (2005) Comment on “Force-clamp spectroscopy monitors the folding trajectory of a single protein”. Science 308:498bGoogle Scholar
  14. Bilsel O, Matthews CR (2006) Molecular dimensions and their distributions in early folding intermediates. Curr Opin Struct Biol 16:86–93PubMedGoogle Scholar
  15. Binnig G, Rohrer H (1986) Scanning tunneling microscopy. IBM J Res Devel 30:355–369Google Scholar
  16. Binnig G, Quate CF, Gerber C (1986) Atomic force microscope. Phys Rev Lett 56:930–933PubMedGoogle Scholar
  17. Bornschlögl T, Rief M (2008) Single-Molecule dynamics of mechanical coiled-coil unzipping. Langmuir 24:1338–1342PubMedGoogle Scholar
  18. Bornschlögl T, Gebhardt JCM, Rief M (2009a) Designing the folding mechanics of coiled coils. Chemphyschem 10:2800–2804PubMedGoogle Scholar
  19. Bornschlögl T, Anstrom DM, Mey E, Dzubiella J, Rief M, Forest KT (2009b) Tightening the knot in phytochrome by single-molecule atomic force microscopy. Biophys J 96:1508–1514PubMedGoogle Scholar
  20. Brandenburg B, Zhuang X (2007) Virus trafficking-learning from single-virus tracking. Nat Rev Microbiol 5:197–208PubMedGoogle Scholar
  21. Brucale M, Sandal M, Di Maio S, Rampioni A, Tessari I, Tosatto L, Bisaglia M, Bubacco L, Samorì B (2009) Pathogenic mutations shift the equilibria of α-synuclein single molecules towards structured conformers. Chembiochem 10:176–183PubMedGoogle Scholar
  22. Brujic J, Fernandez JM (2005) Response to comment on “Force-clamp spectroscopy monitors the folding trajectory of a single protein”. Science 308:498cGoogle Scholar
  23. Bryngelson JD, Onuchic JN, Socci ND, Wolynes PG (1995) Funnels, pathways, and the energy landscape of protein folding: a synthesis. Proteins 21:167–195PubMedGoogle Scholar
  24. Bullard B, Garcia T, Benes V, Leake MC, Linke WA, Oberhauser AF (2006) The molecular elasticity of the insect flight muscle proteins projectin and kettin. Proc Natl Acad Sci USA 103:4451–4456PubMedGoogle Scholar
  25. Bustamante C, Chemla Y, Forde N, Izhaky D (2004) Mechanical processes in biochemistry. Annu Rev Biochem 73:705–748PubMedGoogle Scholar
  26. Cao Y, Yoo T, Li H (2008a) Single molecule force spectroscopy reveals engineered metal chelation is a general approach to enhance mechanical stability of proteins. Proc Natl Acad Sci USA 105:11152–11157PubMedGoogle Scholar
  27. Cao Y, Yoo T, Zhuang S, Li H (2008b) Protein–protein interaction regulates proteins mechanical stability. J Mol Biol 378:1132–1141PubMedGoogle Scholar
  28. Cao Y, Er KS, Parhar R, Li H (2009) A force-spectroscopy-based single-molecule metal-binding assay. Chemphyschem 10:1450–1454PubMedGoogle Scholar
  29. Carrión-Vázquez M, Oberhauser AF, Fowler SB, Marszalek PE, Broedel SE, Clarke J, Fernandez JM (1999) Mechanical and chemical unfolding of a single protein: a comparison. Proc Natl Acad Sci USA 96:3694–3699PubMedGoogle Scholar
  30. Carrión-Vázquez M, Oberhauser AF, Fisher TE, Marszalek PE, Li H, Fernandez JM (2000) Mechanical design of proteins studied by single-molecule force spectroscopy and protein engineering. Prog Biophys Mol Biol 74:63–91PubMedGoogle Scholar
  31. Carrión-Vázquez M, Li H, Lu H, Marszalek PE, Oberhauser AF, Fernandez JM (2003) The mechanical stability of ubiquitin is linkage dependent. Nat Struct Biol 10:738–743PubMedGoogle Scholar
  32. Carrión-Vázquez M, Oberhauser AF, Díez H, Hervás R, Oroz J, Fernández J, Marínez-Martín D (2006) Protein nanomechanics—as studied by AFM single-molecule force spectroscopy. In: Arrondo JLR, Alonso A (eds) Advanced techniques in biophysics. Springer, Heidelberg, pp 163–245Google Scholar
  33. Carrión-Vázquez M, Cieplak M, Oberhauser AF (2009) Protein mechanics at the single-molecule level. In: Meyers RA (ed) Encyclopedia of complexity and systems science. Springer, New York, NY, pp 7026–7051Google Scholar
  34. Cieplak M, Szymczak P (2006) Protein folding in a force clamp. J Chem Phys 124:194901–194904PubMedGoogle Scholar
  35. Cox BS (1965) [PSI+], a cytoplasmic suppressor of super-suppressors in yeast. Heredity 20:505–521Google Scholar
  36. Crick SL, Jayaraman M, Frieden C, Wetzel R, Pappu RV (2006) Fluorescence correlation spectroscopy shows that monomeric polyglutamine molecules form collapsed structures in aqueous solutions. Proc Natl Acad Sci USA 103:16764–16769PubMedGoogle Scholar
  37. Cummings CJ, Zoghbi HY (2000) Trinucleotide repeats: mechanisms and pathophysiology. Annu Rev Genomics Hum Genet 1:281–328PubMedGoogle Scholar
  38. Dembo M, Torney DC, Saxman K, Hammer D (1988) The reaction-limited kinetics of membrane-to-surface adhesion and detachment. Proc R Soc Lond B Biol Sci 234:55–83PubMedGoogle Scholar
  39. Deniz AA, Laurence TA, Beligere GS, Dahan M, Martin AB, Chemla DS, Dawson PE, Schultz PG, Weiss S (2000) Single-molecule protein folding: diffusion fluorescence resonance energy transfer studies of the denaturation of chymotrypsin inhibitor 2. Proc Natl Acad Sci USA 97:5179–5184PubMedGoogle Scholar
  40. Deniz AA, Laurence TA, Dahan M, Chemla DS, Schultz PG, Weiss S (2001) Ratiometric single-molecule studies of freely diffusing biomolecules. Annu Rev Phys Chem 52:233–253PubMedGoogle Scholar
  41. Dougan L, Li J, Badilla CL, Berne BJ, Fernandez JM (2009) Single homopolypeptide chains collapse into mechanically rigid conformations. Proc Natl Acad Sci USA 106:12605–12610PubMedGoogle Scholar
  42. Dunker AK, Brown CJ, Lawson JD, Iakoucheva LM, Obradovic Z (2000a) Intrinsic disorder and protein function. Biochemistry 41:6573–6582Google Scholar
  43. Dunker AK, Obradovic Z, Romero P, Garner EC, Brown CJ (2000b) Intrinsic protein disorder in complete genomes. Genome Inform Ser Workshop Genome Inform 11:161–171PubMedGoogle Scholar
  44. Dunker AK, Lawson JD, Brown CJ, Williams RM, Romero P, Oh JS, Oldfield CJ, Campen AM, Ratliff CM, Hipps KW, Ausio J, Nissen MS, Reeves R, Kang CH, Kissinger CR, Bailey RW, Griswold MD, Chiu W, Garner EC, Obradovic Z (2001) Intrinsically disordered protein. J Mol Graph Model 19:26–59PubMedGoogle Scholar
  45. Evans E, Ritchie K (1997) Dynamic strength of molecular adhesion bonds. Biophys J 72:1541–1555PubMedGoogle Scholar
  46. Faux NG, Bottomley SP, Lesk AM, Irving JA, Morrison JR, de la Banda MG, Whisstock JC (2005) Functional insights from the distribution and role of homopeptide repeat-containing proteins. Genome Res 15:537–551PubMedGoogle Scholar
  47. Fernandez JM, Li H (2004) Force-clamp spectroscopy monitors the folding trajectory of a single protein. Science 303:1674–1678PubMedGoogle Scholar
  48. Fernandez JM, Li H, Brujic J (2004) Response to comment on “Force-clamp spectroscopy monitors the folding trajectory of a single protein”. Science 306:411cGoogle Scholar
  49. Ferreon AC, Moran CR, Gambin Y, Deniz AA (2010) Single-molecule fluorescence studies of intrinsically disordered proteins. Methods Enzymol 472:179–204PubMedGoogle Scholar
  50. Forman JR, Yew ZT, Qamar S, Sandford RN, Paci E, Clarke J (2009) Non-native interactions are critical for mechanical strength in PKD domains. Structure 17:1582–1590PubMedGoogle Scholar
  51. Galera-Prat A, Gómez-Sicilia A, Oberhauser AF, Cieplak M, Carrión-Vázquez M (2010) Understanding biology by stretching proteins: recent progress. Curr Opin Struct Biol 20:63–69PubMedGoogle Scholar
  52. Garcia-Manyes S, Dougan L, Badilla CL, Brujic J, Fernandez JM (2009a) Direct observation of an ensemble of stable collapsed states in the mechanical folding of ubiquitin. Proc Natl Acad Sci USA 106:10534–10539PubMedGoogle Scholar
  53. Garcia-Manyes S, Liang J, Szoszkiewicz KTL, Fernández JM (2009b) Force-activated reactivity switch in a bimolecular chemical reaction. Nat Chem 1:236–242PubMedGoogle Scholar
  54. Gebhardt JCM, Bornsclögl T, Rief M (2010) Full distance-resolved folding energy landscape of one single protein molecule. Proc Natl Acad Sci USA 107:2013–2018PubMedGoogle Scholar
  55. Gräter F, Shen J, Jiang H, Gautel M, Grubmüller H (2005) Mechanicallyinduced titin kinase activation studied by force-probe molecular dynamicssimulations. Biophys J 88:790–804PubMedGoogle Scholar
  56. Grützner A, Garcia-Manyes S, Kotter S, Badilla CL, Fernandez JM, Linke WA (2009) Modulation of titin-based stiffness by disulfide bonding in the cardiac titin N2-B unique sequence. Biophys J 97:825–834PubMedGoogle Scholar
  57. Guzmán DL, Randall A, Baldi P, Guan Z (2010) Computational and single-molecule force studies of a macro domain protein reveal a key molecular determinant for mechanical stability. Proc Natl Acad Sci USA 107:1989–1994PubMedGoogle Scholar
  58. Helmes M, Trombitás K, Centner T, Kellermayer M, Labeit S, Linke WA, Granzier H (1999) Mechanically driven contour-length adjustment in rat cardiac titin’s unique N2B sequence: titin is an adjustable spring. Circ Res 84:1339–1352PubMedGoogle Scholar
  59. Hervás R, Oroz J, Galera-Prat A, Goñi O, Valbuena A, Vera AM, Gómez-Sicilia A, Losada-Urzáiz F, Uversky VN, Menéndez M, Laurents DV, Bruix M, Carrión-Vázquez M (2012) Common features at the start of the neurodegeneration cascade. PLoS Biol 10(5):e1001335. doi:10.1371/journal.pbio.10011335 PubMedGoogle Scholar
  60. Hirokawa N, Shiomura Y, Okabe S (1988) Tau proteins: the molecular structure and mode of binding on microtubules. J Cell Biol 107:1449–1459PubMedGoogle Scholar
  61. Humphrey W, Dalke A, Schulten K (1996) VMD: visual molecular dynamics. J Mol Graph 14:33–38PubMedGoogle Scholar
  62. Ishijima A, Kojima H, Funatsu T, Tokunaga M, Higuchi H, Tanaka H, Yanagida T (1998) Simultaneous observation of individual ATPase and mechanical events by a single myosin molecule during interaction with actin. Cell 92:161–171PubMedGoogle Scholar
  63. Jahn TR, Parker MJ, Homans SW, Radford SE (2006) Amyloid formation under physiological conditions proceeds via a native-like folding intermediate. Nat Struct Mol Biol 13:195–201PubMedGoogle Scholar
  64. Johnson CP, Tang HY, Carag C, Speicher DW, Discher DE (2007) Forced unfolding of proteins within cells. Science 317:663–666PubMedGoogle Scholar
  65. Jolleymore A, Li H (2010) Measuring “unmeasurable” folding kinetics of proteins by single-molecule force spectroscopy. J Mol Biol 402:610–617Google Scholar
  66. Junker JP, Rief M (2009) Single-molecule force spectroscopy distinguishes target binding modes of calmodulin. Proc Natl Acad Sci USA 106:14361–14366PubMedGoogle Scholar
  67. Junker JP, Hell K, Schlierf M, Neupert W, Rief M (2005) Influence of substrate binding on the mechanical stability of mouse dihydrofolate reductase. Biophys J 89:45–48Google Scholar
  68. Junker JP, Ziegler F, Rief M (2009) Ligand-dependent equilibrium fluctuations of single calmodulin molecules. Science 323:633–637PubMedGoogle Scholar
  69. Kazmierczak P, Müller U (2012) Sensing sound: molecules that orchestrate mechanotransduction by hair cells. Trends Neurosci 35:220–229PubMedGoogle Scholar
  70. Khare SD, Ding F, Gwanmesia KN, Dokholyan NV (2005) Molecular origin of polyglutamine aggregation in neurodegenerative diseases. PLoS Comput Biol 1:230–235PubMedGoogle Scholar
  71. Kidd M (1963) Paired helical filaments in electron microscopy of Alzheimer’s disease. Nature 197:192–193PubMedGoogle Scholar
  72. Kim HY, Heise H, Fernandez CO, Baldus M, Zweckstetter M (2007) Correlation of amyloid fibril β-structure with the unfolded state of α-synuclein. Chembiochem 8:1671–1674PubMedGoogle Scholar
  73. King CY, Tittmann P, Gross H, Gebert R, Aebi M, Wüthrich K (1997) Prion-inducing domain 2–114 of yeast Sup35 protein transforms in vitro into amyloid-like filaments. Proc Natl Acad Sci USA 94:6618–6622PubMedGoogle Scholar
  74. Lang MJ, Fordyce PM, Engh AM, Neuman KC, Block SM (2004) Simultaneous, coincident optical trapping and single-molecule fluorescence. Nat Methods 1:133–139PubMedGoogle Scholar
  75. Leake MC, Grutzner A, Kruger M, Linke WA (2006) Mechanical properties of cardiac titin’s N2B-region by single-molecule atomic force spectroscopy. J Struct Biol 155:263–272PubMedGoogle Scholar
  76. Lee JC, Langen R, Hummel PA, Gray HB, Winkler JR (2004) α-synuclein structures from fluorescence energy-transfer kinetics: implications for the role of the protein in Parkinson’s disease. Proc Natl Acad Sci USA 101:16466–16471PubMedGoogle Scholar
  77. Li DY, Brooke B, Davis EC, Mecham RP, Sorensen LK, Boak BB, Eichwald E, Keating MT (1998) Elastin is an essential determinant of arterial morphogenesis. Nature 393:276–280PubMedGoogle Scholar
  78. Li H, Oberhauser AF, Redick SD, Carrion-Vazquez M, Erickson HP, Fernandez JM (2001) Multiple conformations of PEVK proteins detected by single-molecule techniques. Proc Natl Acad Sci USA 98:10682–10686PubMedGoogle Scholar
  79. Li H, Linke WA, Oberhauser AF, Carrion-Vazquez M, Kerkvliet JG, Lu H, Marszalek PE, Fernandez JM (2002) Reverse engineering of the giant muscle protein titin. Nature 418:998–1002PubMedGoogle Scholar
  80. Li H, Wang H-C, Cao Y, Sharma D, Wang M (2008) Configurational entropy modulates the mechanical stability of protein GB1. J Mol Biol 379:871–880PubMedGoogle Scholar
  81. Li J, Fernandez JM, Berne J (2010) Water’s role in the force-induced unfolding of ubiquitin. Proc Natl Acad Sci USA 107:19284–19289PubMedGoogle Scholar
  82. Li PT, Bustamante C, Tinoco I Jr (2007) Real-time control of the energy landscape by force directs the folding of RNA molecules. Proc Natl Acad Sci USA 104:7039–7044PubMedGoogle Scholar
  83. Liang J, Fernandez JM (2011) Kinetic measurements on single-molecule disulfide bond cleavage. J Am Chem Soc 133:3528–3534PubMedGoogle Scholar
  84. Linke WA, Rudy DE, Centner T, Gautel M, Witt C, Labeit S, Gregorio CC (1999) I-band titin in cardiac muscle is a three-element molecular spring and is critical for maintaining thin filament structure. J Cell Biol 146:631–644PubMedGoogle Scholar
  85. Lu H, Schulten K (1999) Steered molecular dynamics simulations of force-induced protein domain unfolding. Proteins 35:453–463PubMedGoogle Scholar
  86. Lu H, Schulten K (2000) The key event in force-induced unfolding of titin’s inmunoglobulin domains. Biophys J 79:51–65PubMedGoogle Scholar
  87. Lu H, Isralewitz B, Krammer A, Vogel V, Schulten K (1998) Unfolding of titin immunoglobulin domains by steered molecular dynamics simulation. Biophys J 75:662–671PubMedGoogle Scholar
  88. Ma L, Xu M, Oberhauser A (2010) Naturally occurring osmolytes modulate the nanomechanical properties of polycystic kidney disease domains. J Biol Chem 285:38438–38443PubMedGoogle Scholar
  89. Marchut AJ, Hall CK (2006) Effects of chain length on the aggregation of model polyglutamine peptides: molecular dynamics simulations. Proteins 66:96–109Google Scholar
  90. Marshall BT, Long M, Piper JW, Yago T, McEver RP, Zhu C (2003) Direct observation of catch bonds involving cell-adhesion molecules. Nature 423:190–193PubMedGoogle Scholar
  91. Marszalek PE, Lu H, Li H, Carrion-Vazquez M, Oberhauser AF, Schulten K, Fernandez JM (1999) Mechanical unfolding intermediates in titin modules. Nature 402:100–103PubMedGoogle Scholar
  92. Martin J (2002) Requirement for GroEL/GroES-dependent protein folding under nonpermissive conditions of macromolecular crowding. Biochemistry 41:5050–5055PubMedGoogle Scholar
  93. Matsunaga Y, Komatsuzaki T (2004) Protein folding dynamics: ergodic behavior in principal component space. Aip Conf Proc 708:342–343Google Scholar
  94. Michalet X, Weiss S, Jager M (2006) Single-molecule fluorescence studies of protein folding and conformational dynamics. Chem Rev 106:1785–1813PubMedGoogle Scholar
  95. Minton AP (2001) The influence of macromolecular crowding and macromolecular confinement on biochemical reactions in physiological media. J Biol Chem 276:10577–10580PubMedGoogle Scholar
  96. Nagai Y, Inui T, Popiel HA, Fujikake N, Hasegawa K, Urade Y, Goto Y, Naiki H, Toda T (2007) A toxic monomeric conformer of the polyglutamine protein. Nat Struct Biol 14:332–340Google Scholar
  97. Neuman KC, Nagy A (2008) Single-moleculeforcespectroscopy: optical tweezers, magnetictweezers and atomic force microscopy. Nat Methods 5:491–505PubMedGoogle Scholar
  98. Ng SP, Rounsevell RWS, Steward A, Geierhaas CD, Williams PM, Paci E, Clarke J (2005) Mechanical unfolding of TNfn3: the unfolding pathway of a fnIII domain probed by protein engineering, AFM and MD simulation. J Mol Biol 350:776–789PubMedGoogle Scholar
  99. Oberhauser AF, Marszalek PE, Carrión-Vázquez M, Fernandez JM (1999) Single protein misfolding events captured by atomic force microscopy. Nat Struct Biol 6:1025–1028PubMedGoogle Scholar
  100. Oroz J, Valbuena A, Vera AM, Mendieta J, Gomez-Puertas P, Carrion-Vazquez M (2011) Nanomechanics of the cadherin ectodomain: canalization by Ca2+ binding results in a new mechanical element. J Biol Chem 286:9405–9418PubMedGoogle Scholar
  101. Oroz J, Hervás R, Carrión-Vázquez M (2012) Unequivocal single-molecule force spectroscopy of proteins by AFM using pFS vectors. Biophys J 102:682–690PubMedGoogle Scholar
  102. Peng Q, Li H (2008) Atomic force microscopy reveals parallel mechanical unfolding pathways of T4 lysozyme: evidence for a kinetic partitioning mechanism. Proc Natl Acad Sci USA 105:1885–1890PubMedGoogle Scholar
  103. Perez-Jimenez R, Li J, Kosuri P, Sanchez-Romero I, Wiita AP, Rodriguez-Larrea D, Chueca A, Holmgren A, Miranda-Vizuete A, Becker K, Cho SH, Beckwith J, Gelhaye E, Jacquot JP, Gaucher EA, Sanchez-Ruiz JM, Berne BJ, Fernandez JM (2009) Diversity of chemical mechanisms in thioredoxin catalysis revealed by single-molecule force-spectroscopy. Nat Struct Mol Biol 16:890–897PubMedGoogle Scholar
  104. Perez-Jimenez R, Inglés-Prieto A, Zhao ZM, Sanchez-Romero I, Alegre-Cebollada J, Kosuri P, Garcia-Manyes S, Kappock TJ, Tanokura M, Holmgren A, Sanchez-Ruiz JM, Gaucher EA, Fernandez JM (2011) Single-molecule paleoenzymology probes the chemistry of resurrected enzymes. Nat Struct Mol Biol 18:592–596PubMedGoogle Scholar
  105. Platt GW, McParland VJ, Kalverda AP, Homans SW, Radford SE (2005) Dynamics in the unfolded state of β 2-microglobulin studied by NMR. J Mol Biol 346:279–294PubMedGoogle Scholar
  106. Popa I, Fernández LM, Garcia-Manyes S (2011) Direct quantification of the attempt frequency determining the mechanical unfolding of ubiquitin protein. J Biol Chem 286:31072–31079PubMedGoogle Scholar
  107. Qian F, Wei W, Germino G, Oberhauser AF (2005) The nanomechanics of polycystin-1 extracellular region. J Biol Chem 280:40723–40730PubMedGoogle Scholar
  108. Rief M, Gautel M, Oesterhelt M, Fernandez JM, Gaub HE (1997) Reversible unfolding of individual titin immunoglobulin domains by AFM. Science 276:1109–1112PubMedGoogle Scholar
  109. Rief M, Pascual J, Saraste M, Gaub HE (1999) Single molecule force spectroscopy of spectrin repeats: low unfolding forces in helix bundles. J Mol Biol 286:553–561PubMedGoogle Scholar
  110. Sandal M, Valle F, Tessari I, Mammi S, Bergantino E, Musiani F, Brucale M, Bubacco L, Samorì B (2008) Conformational equilibria in monomeric α-synuclein at the single-molecule level. PLoS Biol 6:99–108Google Scholar
  111. Sarkar A, Caamano S, Fernandez JM (2005) The elasticity of individual titin PEVK exons measured by single molecule atomic force microscopy. J Biol Chem 280:6261–6264PubMedGoogle Scholar
  112. Sawaya MR, Sambashivan S, Nelson R, Ivanova MI, Sievers SA, Apostol MI, Thompson MJ, Balbirnie M, Wiltzius JJ, McFarlane HT, Madsen AØ, Riekel C, Eisenberg D (2007) Atomic structures of amyloid cross-β spines reveal varied steric zippers. Nature 447:453–457PubMedGoogle Scholar
  113. Schlierf M, Rief M (2005) Temperature softening of a protein in single-molecule experiments. J Mol Biol 354:497–503PubMedGoogle Scholar
  114. Schlierf M, Berkemeier F, Rief M (2007) Direct observation of active protein folding using lock-in force spectroscopy. Biophys J 93:3989–3998PubMedGoogle Scholar
  115. Schuler B (2005) Single-molecule fluorescence spectroscopy of protein folding. Chem Phys Chem 6:1206–1220PubMedGoogle Scholar
  116. Schuler B, Lipman EA, Eaton WA (2002) Probing the free-energy surface for protein folding with single-molecule fluorescence spectroscopy. Nature 419:743–747PubMedGoogle Scholar
  117. Schwaiger I, Schleicher M, Noegel AA, Rief M (2005) The folding pathway of a fast-folding immunoglobulin domain revealed by single-molecule mechanical experiments. EMBO Rep 6:46–51PubMedGoogle Scholar
  118. Sgourakis NG, Yan Y, McCallum SA, Wang C, Garcia AE (2007) The Alzheimer’s peptides Aβ40 and 42 adopt distinct conformations in water: a combined MD/NMR study. J Mol Biol 368:1448–1457PubMedGoogle Scholar
  119. Serquera D, Lee W, Settanni G, Marszalek PE, Paci E, Itzhaki LS (2010) Mechanical unfolding of an ankyrin repeat protein. Biophys J 98:1294–1301PubMedGoogle Scholar
  120. Shankar GM, Li S, Mehta TH, Garcia-Munoz A, Shepardson NE, Smith I, Brett FM, Farrell MA, Rowan MJ, Lemere CA, Regan CM, Walsh DM, Sabatini BL, Selkoe DJ (2008) Amyloid β-protein dimers isolated directly from Alzheimer brains impair synaptic plasticity and memory. Nat Med 14:837–842PubMedGoogle Scholar
  121. Sharma D, Feng G, Khor D, Genchev GZ, Lu H, Li H (2008) Stabilization provided by neighboring strands is critical for the mechanical stability of proteins. Biophys J 95:3935–3942PubMedGoogle Scholar
  122. Sikora M, Cieplak M (2011) Mechanical stability of multidomain proteins and novel mechanical clamps. Proteins 79:1786–1799PubMedGoogle Scholar
  123. Sikora M, Sulowska JI, Cieplak M (2009) Mechanical strength of 17134 model proteins and cystein slipknots. PLoS Comput Biol 5(10):e1000547. doi:10.1371/journal.pcbi.1000547 PubMedGoogle Scholar
  124. Sosnick TR (2004) Comment on “Force-clamp spectroscopy monitors the folding trajectory of a single protein”. Science 306:411bGoogle Scholar
  125. Sotomayor M, Schulten K (2007) Single-molecule experiments in vitro and in silico. Science 316:1144–1148PubMedGoogle Scholar
  126. Still WC, Tempczyk A, Hawley RC, Hendrickson T (1990) Semianalytical treatment of solvation for molecular mechanics and dynamics. J Am Chem Soc 112:6127–6129Google Scholar
  127. Terakawa T, Takada S (2011) Multiscale ensemble modeling of intrinsically disordered proteins: p53 N-terminal domain. Biophys J 101:1450–1458PubMedGoogle Scholar
  128. Tycko R (2011) Solid-state NMR studies of amyloid fibril structure. Annu Rev Phys Chem 62:279–299PubMedGoogle Scholar
  129. Tinnefeld P, Sauer M (2005) Branching out of single-molecule fluorescence spectroscopy: challenges for chemistry and influence on biology. Angew Chem Int Ed 44:2642–2671Google Scholar
  130. Urry DW, Hugel T, Seitz M, Gaub HE, Sheiba L, Dea J, Xu J, Parker T (2002) Elastin: a representative ideal protein elastomer. Philos Trans R Soc Lond B Biol Sci 357:169–184PubMedGoogle Scholar
  131. Uversky VN, Oldfield CJ, Dunker AK (2005) Showing your ID: intrinsic disorder as an ID for recognition, regulation and cell signaling. J Mol Recognit 18:343–384PubMedGoogle Scholar
  132. Uversky VN, Oldfield CJ, Dunker AK (2008) Intrinsically disordered proteins in human diseases: introducing the D2 concept. Annu Rev Biophys Biomol Struct 37:215–246Google Scholar
  133. Uversky VN, Oldfield CJ, Midic U, Xie H, Xue B, Vucetic S, Iakoucheva LM, Obradovic Z, Dunker AK (2009) Unfoldomics of human diseases: linking protein intrinsic disorder with diseases. BMC Genomics 10(Suppl 1):S7PubMedGoogle Scholar
  134. Valbuena A, Oroz J, Hervás R, Vera AM, Rodríguez D, Menéndez M, Sulkowska JI, Cieplak M, Carrión-Vázquez M (2009) On the remarkable mechanostability of scaffoldins and the mechanical clamp motif. Proc Natl Acad Sci USA 106:13791–13796PubMedGoogle Scholar
  135. Valiaev A, Lim DW, Schmidler S, Clark RL, Chilkoti A, Zauscher S (2008) Hydration and conformational mechanics of single, end-tethered elastin-like polypeptides. J Am Chem Soc 130:10939–10946PubMedGoogle Scholar
  136. Vitalis A, Wang X, Pappu RV (2008a) Atomistic simulations of the effects of polyglutamine chain length and solvent quality on conformational equilibria and spontaneous homodimerization. J Mol Biol 384:279–297PubMedGoogle Scholar
  137. Vitalis A, Wang X, Pappu RV (2008b) Quantitative characterization of intrinsic disorder in polyglutamine: insights from analysis based on polymer theories. Biophys J 93:1923–1937Google Scholar
  138. Vogel V, Sheetz MP (2006) Local force and geometry sensing regulate cell functions. Nat Rev Mol Cell Biol 7:265–275PubMedGoogle Scholar
  139. Vogel V, Sheetz MP (2009) Mechanical forces matter in health and disease: from cancer to tissue engineering. In: Vogel V (ed) Nanomedicine. Wiley-VCH, Weinheim, pp 235–303Google Scholar
  140. Wallin S, Zeldovich KB, Shakhnovich EI (2007) Folding mechanics of a knotted protein. J Mol Biol 368:884–893PubMedGoogle Scholar
  141. Walther KA, Gräter F, Dougan L, Badilla CL, Berne BJ, Fernandez JM (2007) Signatures of hydrophobic collapse in extended proteins captured with force spectroscopy. Proc Natl Acad Sci USA 104:7916–7921PubMedGoogle Scholar
  142. Watanabe K, Muhle-Goll C, Kellermayer MS, Labeit S, Granzier H (2002) Molecular mechanics of cardiac titin’s PEVK and N2B spring elements. J Biol Chem 277:11549–11558PubMedGoogle Scholar
  143. Wegmann S, Schöler J, Bippes CA, Mandelkow E, Muller DJ (2011) Competing interactions stabilize pro- and anti-aggregant conformations of human tau. J Biol Chem 286:20512–20524PubMedGoogle Scholar
  144. Wiita AP, Ainavarapu SRK, Huang HH, Fernandez JM (2006) Force-dependent chemical kinetics of disulfide bond reduction observed with single-molecule techniques. Proc Natl Acad Sci USA 103:7222–7227PubMedGoogle Scholar
  145. Wiita AP, Perez-Jimenez R, Walther KA, Gräter F, Berne BJ, Holmgren A, Sanchez-Ruiz JM, Fernandez JM (2007) Probing the chemistry of thioredoxin catalysis with force. Nature 450:124–127PubMedGoogle Scholar
  146. Wright PE, Dyson HJ (1999) Intrinsically unstructured proteins: re-assessing the protein structure-function paradigm. J Mol Biol 293:321–331PubMedGoogle Scholar
  147. Xia F, Thirumalai D, Gräter F (2011) Minimum energy compact structures in force-quench polyubiquitin folding are domain swapped. Proc Natl Acad Sci USA 108:6963–6968PubMedGoogle Scholar
  148. Xie XS, Choi PJ, Li GW, Lee NK, Lia G (2008) Single-molecule approach to molecular biology in living bacterial cells. Annu Rev Biophys 37:417–444PubMedGoogle Scholar
  149. Xiong K, Zwier MC, Myshakina NS, Burger VM, Asher SA, Chong LT (2011) Direct observations of conformational distributions of intrinsically disordered p53 peptides using UV Raman and explicit solvent simulations. J Phys Chem 115:9520–9527Google Scholar
  150. Yamasaki R, Wu Y, McNabb M, Greaser M, Labeit S, Granzier H (2002) Protein kinase A phosphorylates titin’s cardiac-specific N2B domain and reduces passive tension in rat cardiac myocytes. Circ Res 90:1181–1188PubMedGoogle Scholar
  151. Yang M, Teplow D (2008) Amyloid β-protein monomer folding: free-energy surfaces reveal alloform-specific differences. J Mol Biol 384:450–464PubMedGoogle Scholar
  152. Yuan J, Chyan C, Zhou H, Chung T, Peng H, Ping G, Yang G (2008) The effects of macromolecular crowding on the mechanical stability of protein molecules. Protein Sci 17:2156–2166PubMedGoogle Scholar
  153. Zhang S, Iwata K, Lachenmann M, Peng J, Li S, Stimson ER, Lu Y, Felix AM, Maggio JE, Lee JP (2000) The Alzheimer’s peptide Aβ adopts a collapsed coil structure in water. J Struct Biol 130:130–141PubMedGoogle Scholar
  154. Zheng P, Cao Y, Bu T, Straus SK, Li H (2011) Single molecule force spectroscopy reveals that electrostatic interactions affect the mechanical stability of proteins. Biophys J 100:1534–1541PubMedGoogle Scholar
  155. Zhuang S, Linhananta A, Li H (2010) Phenotypic effects of Ehlers–Danlos syndrome-associated mutation on the FnIII domain of tenascin-X. Protein Sci 19:2231–2239PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2012

Authors and Affiliations

  • Rubén Hervás
    • 1
    • 2
  • Albert Galera-Prat
    • 1
    • 2
  • Àngel Gómez-Sicilia
    • 1
    • 2
  • Fernando Losada-Urzáiz
    • 1
    • 2
  • María del Carmen Fernández
    • 1
    • 2
  • Débora Fernández-Bravo
    • 1
    • 2
  • Elena Santana
    • 1
    • 2
  • Clara Barrio-García
    • 1
    • 2
  • Carolina Melero
    • 1
    • 2
  • Mariano Carrión-Vázquez
    • 1
    • 2
  1. 1.Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED)Instituto Cajal, IC-CSICMadridSpain
  2. 2.Instituto Madrileño de Estudios Avanzados en Nanociencia (IMDEA-Nanociencia)MadridSpain

Personalised recommendations