Abstract
RecA fibrils in physiological conditions have been successfully imaged using Tapping Mode atomic force microscopy. This represents the first time images of recA have been obtained without drying, freezing and/or exposure to high vacuum conditions. While previously observed structures – the monomer, the hexamer, the short rod – were seen, a new type of fibril was also observed. This protofibril is narrower in diameter than the standard fibril, and occurs in three distinct morphologies: aperiodic, 100-nm periodic, and 150-nm periodic. In addition, much longer rods were observed, and appear curved and even circular.
Key words: RecA, recombination, assembly, AFM
References
- 1.Roca, A.I. and Cox, M.M.: RecA Protein: Structure, Function, and Role in Recombinational DNA Repair, Prog. Nucleic Acid Res. Mol. Biol.56 (1997), 129–223. [DOI] [PubMed]
- 2.Kodadek, T.: Mechanistic Parallels Between DNA Replication, Recombination and Transcription, Trends Biochem. Sci.23 (1998), 79–83. [DOI] [PubMed]
- 3.Ogawa, T., et al.: Similarity of the yeast RAD51 Filament to the Bacterial RecA Filament, Science259 (1993), 1896–1899. [DOI] [PubMed]
- 4.Rashid, N., Morikawa, M. and Imanaka, T.: A RecA/RAD51 Homologue from a Hyperthermophilic Archaeon Retains the Major RecA Domain Only, Mol. Gen. Genet.253 (1996) 397–400. [DOI] [PubMed]
- 5.Levin-Zaidman, S., et al.: Ordered Intracellular RecA-DNA Assemblies: A Potential Site of in Vivo RecA-mediated Activities, Proc. Natl. Acad. Sci. U. S. A.97 (2000), 6791–6796. [DOI] [PMC free article] [PubMed]
- 6.Story, R.M., Weber, I.T. and Steitz, T.A.: The Structure of the E. coli RecA Protein Monomer and Polymer [Published Erratum appears in Nature 1992 Feb 6;355(6360):567] [see comments], Nature355 (1992), 318–325. [DOI] [PubMed]
- 7.Brenner, S.L., Zlotnick, A. and Griffith, J.D.: RecA Protein Self-Assembly. Multiple Discrete Aggregation States, J. Mol. Biol.204 (1988), 959–972. [DOI] [PubMed]
- 8.Egelman, E.H. and Stasiak, A.: Structure of Helical RecA-DNA Complexes. Complexes Formed in the Presence of ATP-Gamma-S or ATP, J. Mol. Biol.191 (1986), 677–697. [DOI] [PubMed]
- 9.Logan, K.M., et al.: Mutant RecA Proteins which form Hexamer-Sized Oligomers, J. Mol. Biol.266 (1997), 306–316. [DOI] [PubMed]
- 10.Neuendorf, S.K. and Cox, M.M.: Exchange of RecA Protein Between Adjacent RecA Protein-Single-Stranded DNA Complexes, J. Biol. Chem.261 (1986), 8276–8282. [PubMed]
- 11.Williams, R.C. and Spengler, S.J.: Fibers of RecA Protein and Complexes of RecA Protein and Single-Stranded Phi X174 DNA as Visualized by Negative-Stain Electron Microscopy, J. Mol. Biol.187 (1986), 109–118. [DOI] [PubMed]
- 12.Yu, X., et al.: Structural Polymorphism of the RecA Potein from the Thermophilic Bacterium Thermus Aquaticus, Biophys. J.69 (1995), 2728–2738. [DOI] [PMC free article] [PubMed]
- 13.Amrein, M., et al.: Scanning Tunneling Microscopy of RecA-DNA Complexes Coated with a Conducting Film, Science240 (1988), 514–516. [DOI] [PubMed]
- 14.Amrein, M., et al.: Scanning Tunneling Microscopy of Uncoated RecA-DNA Complexes, Science243 (1989), 1708–1711. [DOI] [PubMed]
- 15.Morrical, S.W. and Cox, M.M.: Light Scattering Studies of the RecA Protein of Escherichia Coli: Relationship Between Free RecA Filaments and the RecA X ssDNA Complex, Biochemistry24 (1985), 760–767. [DOI] [PubMed]
- 16.Ellouze, C., et al.: Evidence for Elongation of the Helical Pitch of the RecA Filament upon ATP and ADP Binding using Small-angle Neutron Scattering, Eur. J. Biochem.233 (1995), 579–583. [DOI] [PubMed]
- 17.Wilson, D.H. and Benight, A.S.: Kinetic Analysis of the Pre-equilibrium Steps in the Self-assembly of RecA Protein from Escherichia Coli, J. Biol. Chem.265 (1990), 7351–7359. [PubMed]
- 18.Wittung, P., Norden, B. and Takahashi, M.: Secondary Structure of RecA in Solution. The Effects of Cofactor, DNA and Ionic Conditions, Eur. J. Biochem.228 (1995), 149–154. [DOI] [PubMed]
- 19.Heuser, J. and Griffith, J.: Visualization of RecA Protein and its Complexes with DNA by Quick-freeze/deep-etch Electron Microscopy, J. Mol. Biol.210 (1989), 473–484. [DOI] [PubMed]
- 20.Ruigrok, R.W., et al.: The Inactive form of RecA Protein: The ‘Compact’ Structure, EMBO J, 12 (1993), 9–16. [DOI] [PMC free article] [PubMed]
- 21.Ruigrok, R.W.H. and Dicapua, E.: On the Polymerization State of Reca in the Absence of DNA, Biochimie73 (1991), 191–197. [DOI] [PubMed]
- 22.Timmins, P.A., Ruigrok, R.W.H. and Dicapua, E.: The Solution Structure of RecA Filaments by Small-Angle Neutron-Scattering, Biochimie73 (1991), 227–230. [DOI] [PubMed]
- 23.Budzynski, D.M., Gao, X.H. and Benight, A.S.: Isolation, Characterization, and Magnesium-induced Self-association Kinetics of Discrete Aggregates of RecA Protein from Escherichia Coli, Biopolymers38 (1996), 471–491. [DOI] [PubMed]
- 24.Muller, D.J., et al.: Structural Changes in Native Membrane Proteins Monitored at Subnanometer Resolution with the Atomic Force Microscope: A Review, J. Struct. Biol.119 (1997), 149–157. [DOI] [PubMed]
- 25.Muller, D.J., et al.: Observing Structure, Function and Assembly of Single Proteins by AFM, Prog. Biophys. Mol. Biol.79 (2002), 1–43. [DOI] [PubMed]
- 26.Mou, J.X., et al.: High Resolution Surface Structure of E-coli GroES Oligomer by Atomic Force Microscopy, FEBS Lett.381 (1996), 161–164. [DOI] [PubMed]
- 27.Han, W.H., et al.: Kinked DNA, Nature386 (1997), 563–563. [DOI] [PubMed]
- 28.Vesenka, J., Miller, R. and Henderson, E.: 3-Dimensional Probe Reconstruction for Atomic-force Microscopy, Rev. Sci. Instrum.65 (1994), 2249–2251. [DOI]
- 29.Kuhle, A., et al.: Contrast Artifacts in Tapping Tip Atomic Force Microscopy, Appl. Phys. a-Mater, 66 (1998), S329–S332. [DOI]
- 30.Roca, A.I. and Cox, M.M.: The RecA Protein: Structure and Function, Crit. Rev. Biochem. Mol. Biol.25 (1990), 415–456. [DOI] [PubMed]
- 31.Pollanen, M.S., et al.: Twisted Ribbon Structure of Paired Helical Filaments Revealed by Atomic-Force Microscopy, Am. J. Pathol.144 (1994), 869–873. [PMC free article] [PubMed]
- 32.Sattin, B.D. and Goh, M.C.: Direct Observation of the Assembly of RecA/DNA Complexes by Atomic Force Microscopy, Biophys. J.87 (2004), 3430–3436. [DOI] [PMC free article] [PubMed]
