Revealing the assembly of filamentous proteins with scanning transmission electron microscopy


Autoři: Cristina Martinez-Torres aff001;  Federica Burla aff001;  Celine Alkemade aff001;  Gijsje H. Koenderink aff001
Působiště autorů: Department of Living Matter, AMOLF, Amsterdam, the Netherlands aff001;  Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Faculty of Applied Sciences, Delft University of Technology, Delft, The Netherlands aff002
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article
doi: 10.1371/journal.pone.0226277

Souhrn

Filamentous proteins are responsible for the superior mechanical strength of our cells and tissues. The remarkable mechanical properties of protein filaments are tied to their complex molecular packing structure. However, since these filaments have widths of several to tens of nanometers, it has remained challenging to quantitatively probe their molecular mass density and three-dimensional packing order. Scanning transmission electron microscopy (STEM) is a powerful tool to perform simultaneous mass and morphology measurements on filamentous proteins at high resolution, but its applicability has been greatly limited by the lack of automated image processing methods. Here, we demonstrate a semi-automated tracking algorithm that is capable of analyzing the molecular packing density of intra- and extracellular protein filaments over a broad mass range from STEM images. We prove the wide applicability of the technique by analyzing the mass densities of two cytoskeletal proteins (actin and microtubules) and of the main protein in the extracellular matrix, collagen. The high-throughput and spatial resolution of our approach allow us to quantify the internal packing of these filaments and their polymorphism by correlating mass and morphology information. Moreover, we are able to identify periodic mass variations in collagen fibrils that reveal details of their axially ordered longitudinal self-assembly. STEM-based mass mapping coupled with our tracking algorithm is therefore a powerful technique in the characterization of a wide range of biological and synthetic filaments.

Klíčová slova:

Atomic force microscopy – Collagens – Extracellular matrix proteins – Microtubules – Molecular structure – Packing density – Protein structure – Tobacco mosaic virus


Zdroje

1. Buehler MJ, Yung YC. Deformation and failure of protein materials in physiologically extreme conditions and disease. Nat Mater. 2009; 8: 175–188. doi: 10.1038/nmat2387 19229265

2. Schwarz US, Safran SA. Physics of adherent cells. Rev Mod Phys. 2013; 85: 1327–1381.

3. Burla F, Mulla Y, Vos BE, Aufderhorst-Roberts A, Koenderink GH. From mechanical resilience to active material properties in biopolymer networks. Nat Rev Phys. 2019; 1: 249–263.

4. Wall JS, Hainfeld JF. Mass mapping with the scanning transmission electron microscope. Annu Rev Biophys Biophys Chem. 1986; 15: 355–376. doi: 10.1146/annurev.bb.15.060186.002035 3521658

5. Muller SA, Engel A. Structure and mass analysis by scanning transmission electron microscopy. Micron. 2001; 32: 21–31. doi: 10.1016/s0968-4328(00)00022-6 10900377

6. Engel A. Molecular weight determination by scanning transmission electron microscopy. Ultramicroscopy. 1978; 3: 273–281 doi: 10.1016/s0304-3991(78)80037-0 734784

7. Muller SA, Goldie KN, Burki R, Haring R, Engel A. Factors influencing the precision of quantitative scanning transmission electron microscopy. Ultramicroscopy. 1992; 46: 317–336.

8. Freeman R, Leonard KR. Comparative mass measurement of biological macromolecules by scanning transmission electron microscopy. J Microsc. 1981; 122: 275–86. doi: 10.1111/j.1365-2818.1981.tb01267.x 6113286

9. Bullitt ESA, DeRosier DJ, Coluccio LM, Tilney LG. Three-dimensional reconstruction of an actin bundle. J Cell Biol. 1988; 107: 597–611. doi: 10.1083/jcb.107.2.597 3417764

10. Nojima D, Linck RW, Egelman EH. At least one of the protofilaments in flagellar microtubules is not composed of tubulin. Curr Biol. 1995; 5: 158–167. doi: 10.1016/s0960-9822(95)00037-6 7743179

11. Holmes DF, Graham HK, Trotter JA, Kadler KE. STEM/TEM studies of collagen fibril assembly. Micron. 2001; 32: 273–285. doi: 10.1016/s0968-4328(00)00040-8 11006507

12. Tonino P, Simon M, Craig R. Mass determination of native smooth muscle myosin filaments by scanning transmission electron microscopy. J Mol Biol. 2002; 318: 999–1007. doi: 10.1016/S0022-2836(02)00191-2 12054797

13. Goldsbury C, Baxa U, Simon MN, Steven AC, Engel A, Wall JS, et al. Amyloid structure and assembly: insights from scanning transmission electron microscopy. J Struct Biol. 2011; 173: 1–13. doi: 10.1016/j.jsb.2010.09.018 20868754

14. Krzyzanek V, Mueller SA, Engel A, Reichelt R. MASSDET–A fast and user-firendly multiplatform software for mass determination by dark-field electron microscopy. J Struct Biol. 2009; 165: 78–87 doi: 10.1016/j.jsb.2008.10.006 19041401

15. Bozec L, van der Heijden G, Horton M. Collagen fibrils: nanoscale ropes. Biophys J. 2007; 92: 70–75. doi: 10.1529/biophysj.106.085704 17028135

16. Canny J. A computational approach to edge detection. IEEE Trans Pattern Anal Mach Intell. 1986; 8: 679–968. 21869365

17. Mallat S, Hwang WL. Singularity detection and processing with wavelets. IEEE Trans Inf Theory. 1992; 38: 617–643.

18. Martinez-Torres C, Laperrousaz B, Berguiga L, Boyer-Provera E, Elezgaray J, Nicolini FE, et al. Deciphering the internal complexity of living cells with quantitative phase microscopy: a multiscale approach. J Biomed Opt. 2015; 20: 096005. doi: 10.1117/1.JBO.20.9.096005 26334978

19. Namba K, Stubbs G. Structure of tobacco mosaic virus at 3.6 A resolution: implications for assembly. Science. 1986; 231: 1401–1406 doi: 10.1126/science.3952490 3952490

20. Zimmerman K, Hagedorn H, Heuck CC, Hinrichsen M, Ludwig H. The ionic properties of the filamentous bacteriophages Pf1 and Fd*. J Biol Chem. 1986; 261: 1653–1655. 3944103

21. Merino F, Pospich S, Funk J, Wagner T, Kullmer F, Amdt HD, et al. Structural transitions of F-actin upon ATP hydrolysis at near-atomic resolution revealed by cryo-EM. Nat Struct Mol Biol. 2018; 25: 528–537. doi: 10.1038/s41594-018-0074-0 29867215

22. Chou SZ, Pollard TD. Mechanism of actin polymerization revealed by cryo-EM structures of actin filaments with three different bound nucleotides. Proc Natl Acad Sci U S A. 2019; 13: 201807028

23. Sousa AA, Leapman RD. Development and application of STEM for the biological sciences. Ultramicroscopy. 2012; 126: 38–49.

24. Elbaum M. Quantitative cryo-scanning transmission electron microscopy of biological materials. Adv Mater. 2018; 30: e1706681. doi: 10.1002/adma.201706681 29748979

25. Meurer-Grob P, Kasparian J, Wade H. Microtubule structure at improved resolution. Biochemistry. 2001; 40: 8000–8008. doi: 10.1021/bi010343p 11434769

26. Alushin GM, Lander GC, Kellog EH, Zhang R, Baker D, Nogales E. High-resolution microtubule structures reveal the structural transitions in Αβ-tubulin upon GTP hydrolysis. Cell. 2014; 157: 1117–29. doi: 10.1016/j.cell.2014.03.053 24855948

27. Hoenger A, Thormahlen M, Diaz-Avalos R, Doerhoefer M, Goldie KN, Muller J, et al. New look at the microtubule binding patterns of dimeric kinesins. J Mol Biol. 2000; 297: 1087–1103. doi: 10.1006/jmbi.2000.3627 10764575

28. Holmes DF, Lu Y, Starborg T, Kadler KE. Collagen fibril assembly and function. Curr Top Dev Biol. 2018; 130: 107–142. doi: 10.1016/bs.ctdb.2018.02.004 29853175

29. Hulmes DJ, Wess TJ, Prockop DJ, Fratzl P. Radial packing order and disorder in collagen fibrils. Biophys J. 1995; 68: 1661–1670. doi: 10.1016/S0006-3495(95)80391-7 7612808

30. Zhu J, Kaufman J. Collagen I self-assembly: revealing the developing structures that generate turbidity. Biophys J. 2014; 106: 1822–1831. doi: 10.1016/j.bpj.2014.03.011 24739181

31. Orgel J, Irving T, Miller A, Wess T. Microfibrillar structure of type I collagen In Situ. Proc Natl Academ Sci U S A. 2006; 103: 9001–9005.

32. Fratzl P., Ed. Collagen: structure and mechanics. Springer US; 2008.

33. Kadler KE, Holmes DF, Trotter JA, Chapman JA. Collagen fibril formation. Biochem J. 1996; 316: 1–11. doi: 10.1042/bj3160001 8645190

34. Fraser RD, MacRae TP, Miller A. Molecular packing in type I collagen fibrils. J Mol Biol. 1987, 193: 115–125. doi: 10.1016/0022-2836(87)90631-0 3586015

35. Cameron S, Kreplak L, Ruttenberg AD. Polymorphism of stable collagen fibrils. Soft Matter. 2018; 14: 4772–4783 doi: 10.1039/c8sm00377g 29799597

36. Brown AI, Kreplak L, Rutenber AD. An equilibrium double-twist model for the radial structure of collagen fibrils. Soft Matter. 2014; 10: 8500–11. doi: 10.1039/c4sm01359j 25238208

37. Holmes DF, Chapman JA, Prockop DJ, Kadler KE. Growing tips of type I collagen fibrils formed in vitro are near-paraboloidal in shape, implying a reciprocal relationship between accretion and diameter. Proc Natl Acad Sci U S A. 1992; 89: 9855–9856 doi: 10.1073/pnas.89.20.9855 1409712

38. Ferri F, Calegari GR, Molteni M, Cardinali B, Magatti D, Rocco M. Size and density of fibers in fibrin and other filamentous networks from turbidimetry: beyond a revisited Carr-Hermans method, accounting for fractality and porosity. Macromolecules. 2015; 48: 5423–5432.

39. Abass A, Bell JS, Spang A, Hayes S, Meek KM, Boote C. SAXS4COLL: an integrated software tool for analysing fibrous collagen-based tissues. J Appl Crystallogr. 2017; 50: 1235–1240. doi: 10.1107/S1600576717007877 28808439

40. Misof K, Rapp G, Fratzl P. A new molecular model for collagen elasticity based on synchrotron X-ray scattering evidence. Biophys J. 1997; 72: 1376–81. doi: 10.1016/S0006-3495(97)78783-6 9138582

41. Amenabar I, Poly S, Nuansing W, Hubrich EH, Govyadinov AA, Huth F, Krutokhvostov R, Zhang L, Knez M, Heberle J, Bittner AM, Hillenbrand R,. Structural analysis and mapping of individual protein complexes by infrared nanospectroscopy. Nat Commun. 2013; 4:2890. doi: 10.1038/ncomms3890 24301518

42. Kurouski D, Deckert-Gaudig T, Deckert V, Ledney IK. Structure and composition of insulin fibril surfaces probed by TERS. J Am Chem Soc. 2012; 134: 13323–13329. doi: 10.1021/ja303263y 22813355

43. vandenAkker CC, Deckert-Gaudig T, Schleeger M, Velikov KP, Deckert V, Bonn M, Koenderink GH. Nanoscale heterogeneity of the molecular structure of individual hIAPP amyloid fibrils revealed with tip-enhanced Raman spectroscopy. Small. 2015; 11:4131–9. doi: 10.1002/smll.201500562 25952953

44. Deckert-Gaudig T, Kurouski D, Hedegaard MA, Singh P, Ledney IK, Deckert V. Spatially resolved spectroscopic differentiation of hydrophilic and hydrophobic domains on individual amyloid fibrils. Sci Rep. 2016; 6: 33575. doi: 10.1038/srep33575 27650589

45. Buehler M. Nature designs though collagen: explaining the nanostructure of collagen fibrils. Proc Natl Acad Sci U S A. 2006; 106: 12285–12290.

46. Perimal S, Antipova O, Orgel J. Collagen fibril architecture, domain organization, and triple-helical conformation govern its proteolysis. Proc Natl Acad Sci U S A. 2008; 105: 2824–2829. doi: 10.1073/pnas.0710588105 18287018

47. Brown AE, Litvinov RI, Discher DE, Purohit PK, Weisel JW. Multiscale mechanics of fibrin polymer: gel stretching with protein unfolding and loss of water. Science. 2009; 325: 741–4 doi: 10.1126/science.1172484 19661428

48. Lopez CG, Saldanha O, Aufderhorst-Roberts A, Martinez-Torres C, Kuijs M, Koenderink GH, Köster S, Huber K. Effect of ionic strength on the structure and elongational kinetics of vimentin filaments. Soft Matter. 2018; 14: 8445–8454. doi: 10.1039/c8sm01007b 30191240

49. Protopopova AD, Litvinov RI, Galanakis DK, Nagaswami C, Barinov NA, Mukhitov AR, Klinov DV, Weisel JW. Morphometric characterization of fibrinogen’s αC regions and their role in fibrin self-assembly and molecular organization. Nanoscale. 2017; 9: 13707–13716. doi: 10.1039/c7nr04413e 28884176

50. Kouwer PH, Koepf M, Le Sage VA, Jaspers M, van Buul AM, Eksteen-Akeroyd ZH, et al. Responsive biomimetic networks from polyisocyanopeptide hydrogels. Nature. 2013; 493: 651–5 doi: 10.1038/nature11839 23354048

51. Schuldt C, Schnauß J, Händler T, Glaser M, Lorenz J, Golde T, et al. Tuning synthetic semiflexible networks by bending stiffness. Phys Rev Lett. 2016; 117: 197801. doi: 10.1103/PhysRevLett.117.197801 27858441

52. Knowles TP, Mezzenga R. Amyloid fibrils as building blocks for natural and artificial functional materials. Adv Mater. 2016; 28: 6546–61. doi: 10.1002/adma.201505961 27165397

53. Alvarado J, Koenderink GH. Reconstituting Cytoskeletal Contraction Events with biomimetic actin-myosin active gels. In: Ross J, Marshall W. editors. Building a cell from its component parts. Elsevier: Methods in Cell Biology 128; Amsterdam, 2015. pp. 83–103.

54. Preciado Lopez M, Huber F, Grigoriev I, Steinmetz MO, Akhmanova A, Dogterom M, et al. In Vitro reconstitution of dynamic microtubules interacting with actin filament networks. Methods Enzymol. 2014; 540: 301–20. doi: 10.1016/B978-0-12-397924-7.00017-0 24630114


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2019 Číslo 12