Dwight L. Anderson

Professor Emeritus



Bacteriophage phi29 structure and assembly

Research Summary


Structure and Assembly of Bacteriophage ø29 Bacteriophage ø29 of Bacillus subtilis offers the opportunity for a complete understanding of the structure and assembly of a complex dsDNA bacterial virus in molecular terms. Current goals include uncovering the mechanisms of a) assembly of the viral precursor capsid (prohead) in vitro from purified proteins and a host chaperonin, b) assembly and function of the powerful motor that packages the dsDNA and the mechanochemical energy transduction, c) sequential protein and DNA conformational changes that regulate assembly, and d) tail penetration of the cell wall during infection. Atomic resolution structures of five of eight ø29 structural proteins have been produced, and a complete atomic resolution structure of the virus is within reach. A sequence of single-particle, asymmetric cryoEM-3D reconstructions of mature and empty virions have been produced in collaborations. A sub-nanometer reconstruction has revealed protein secondary structural elements in situ, rearrangement of connector alpha helices, remodeling of the shell protein where it contacts DNA, and a 60Å diameter DNA toroid in the junction between connector/lower collar that may form as the DNA responds to linear compression. The mechanochemical cycle of the packaging motor has been probed by the use of laser tweezers single molecule studies in collaboration. Phosphate or ADP release is likely the power stroke of the motor. Use of optical tweezers capable of 1 bp resolution has revealed a highly coordinated motor in which the gp16 ATPase subunits load with ATP during a “dwell”, followed by rapid firings of the subunits to package 10 bp of DNA in a “burst” which consists, surprisingly, of four 2.5-bp steps. Load bearing contacts with phosphates that regulate the chemical cycle take place during the dwell phase every 10-bp on the 5’-3’ strand in the direction of packaging. Strong phosphate contacts hold the DNA in place during the dwell phase, while less specific contacts drive translocation during the burst phase. The ø29 tail proteins gp13 and gp12* have enzymatic activities, forecasting an understanding of tail penetration of the cell wall in infection. The morphogenetic factor gp13, located at the distal tip of the tail knob, is a Zn2+-D,D-endopeptidase that cleaves the peptide cross-link of the B. subtilis cell wall. Its structure was solved to 1.8Å resolution by X-ray crystallography. An autocatalytic assembly mechanism of the gp12 tail appendages was demonstrated by X-ray crystallography. The gp12 C-terminal domain is an “auto-chaperone” that aids trimerization, and auto-cleavage of the domain is followed by a unique ATP-dependent release. Subsequently, three domains of the N-terminus function in appendage attachment, cell wall digestion, and host binding, respectively.


  • Ding, F., C. Lu, W. Zhao, R. R. Kanagalaghatta, D. L. Anderson, P. J. Jardine, S. Grimes, and A. Ke. 2011. Structure and assembly of the essential RNA ring component of a viral DNA packaging motor. Proc. Natl. Acad. Sci. U.S.A.108:7357-7362.
  • Aathavan, K., A. T. Politzer, A. Kaplan, J. R. Moffitt, Y. R. Chemla, S. Grimes, P. J. Jardine, D. L. Anderson, and C. Bustamante. 2009. Substrate Interactions and Promiscuity in a Viral DNA Packaging Motor. Nature 461:669-673.
  • Xiang, Y., P. G. Leiman, L. Li, S. Grimes, D. L. Anderson, and M. G. Rossmann. 2009. Crystallographic insights into the autocatalytic assembly mechanism of a bacteriophage tail spike. Mol. Cell 34:375-386. PMC Journal - In Process.
  • Cohen, D. N., Y. Y. Sham, G. D. Haugstad, Y. Xiang, M. G. Rossmann, D. L. Anderson, and D. L. Popham. 2009. Shared catalysis in virus entry and bacterial cell wall depolymerization. J. Mol. Biol. 387:607-618. PMC Journal – In Process.
  • Moffitt, J. R., Y. R. Chemla, K. Aathavan, S. Grimes, P. J. Jardine, D. L Anderson, and C. Bustamante. 2009. Intersubunit coordination in a homomeric ring ATPase. Nature 457:446-450. PMC Journal - In Process.
  • Zhao, W., M. C. Morais, D. L. Anderson, P. J. Jardine, and S. Grimes. 2008. Role of the CCA bulge of prohead RNA of bacteriophage ø29 in DNA packaging. J. Mol. Biol. 383:520-528. PMC Journal - In Process.
  • Koti, J. S., M. C. Morais, R. Rajagopal, B. A. L. Owen, C. T. McMurray, and D. L. Anderson. 2008. DNA packaging motor assembly intermediate of bacteriophage ø29. J. Mol. Biol. 381:1114-1132. PMC Journal – In Process.
  • Xiang, Y., M. C. Morais, D. C. Cohen, V. D. Bowman, D. L. Anderson, and M. G. Rossmann. 2008. Crystal and cryoEM structural studies of a cell wall degrading enzyme in the bacteriophage ø29 tail. Proc. Natl. Acad. Sci. U.S.A. 105: 9552-9557. PMCID:PMC2474476.
  • Morais, M. C., J. S. Koti, V. D. Bowman, E. Reyes-Aldrete, D. L. Anderson, and M. G. Rossmann. 2008. Defining molecular and domain boundaries in the bacteriophage ø29 DNA packaging motor. Structure. 16:1267-1274. PMC Journal – In Process.
  • Kitamura, A., P. J. Jardine, D. L. Anderson, S. Grimes, and H. Matsuo. 2008. Analysis of intermolecular base pair formation of prohead RNA of the phage ø29 DNA packaging motor using NMR spectroscopy. Nucleic Acids Res. 36:839-848. PMCID: PMC2241910
  • Cohen, D. N., S. E. Erickson, Y. Xiang, M. G. Rossmann, and  D. L. Anderson. 2008. Multifunctional roles of a bacteriophage phi 29 morphogenetic factor in assembly and infection. J. Mol. Biol. 378: 804-817. PMCID: PMC2443984.
  • Tang, J., N. Olson, P. J. Jardine, S. Grimes, D. L. Anderson, and T. S. Baker. 2008. DNA poised for release in bacteriophage ø29. Structure 6:935-943. PMCID: PMC2556148.
  • Comolli, L. R., A. J. Spakowitz, C. E. Siegerist, P. J. Jardine, S. Grimes, D. L. Anderson, C. Bustamante, and K. H. Downing. 2008. Three-dimensional architecture of the bacteriophage ø29 packaged genome and elucidation of its packaging process. Virology 371:267-277.
  • Rickgauer, J. P., D. N. Fuller, S. Grimes, P. J. Jardine, D. L. Anderson, and D. E. Smith. 2008. Portal motor velocity and internal force resisting viral DNA packaging in bacteriophage ø29. Biophys J. 94:159-167. PMCID: PMC2134861.
  • Fuller, D. N., D. M. Raymer, J. P. Rickgauer, R. M. Robertson, C. E. Catalano, D. L. Anderson, S. Grimes, and D. E. Smith. 2007. Measurements of single DNA molecule packaging dynamics in phage lambda reveal high forces, high motor processivity, and capsid transformations. J. Mol. Biol. 373:1113-1122.
  • Fuller, D., J. P. Rickgauer, P. J. Jardine, S. Grimes, D. L. Anderson, and D. E. Smith. 2007. Ionic effects on viral DNA packaging and portal motor function in bacteriophage ø29. Proc. Natl. Acad. Sci. U. S. A. 104:11245-11250. PMCID: PMC2040884.
  • Atz, R., S. Ma, J. Gao, D. L. Anderson, and S. Grimes. 2007. Alanine scanning and Fe-BABE probing of the bacteriophage ø29 prohead RNA-connector interaction. J. Mol. Biol. 369:239-248. PMCID: PMC1976407
  • Hugel, T., J. Michaelis, C. L. Hetherington, P. J. Jardine, S. Grimes, J. M. Walter, W. Falk, D. L. Anderson, and C. Bustamante. 2007. Experimental test of connector rotation during DNA packaging into bacteriophage ø29 capsids. PLoS Biol. 5:e59. PMCID: PMC1800307.
  • Xiang, Y., M. C. Morais, A. J. Battisti, S. Grimes, P. J. Jardine, D. L. Anderson, and M. G. Rossmann. 2006. Structural changes of bacteriophage ø29 upon DNA packaging and release. EMBO J. 25:5229-5239. PMCID: PMC1630414.
  • Jardine, P. J. and D. Anderson. 2006. DNA packaging in double-stranded DNA bacteriophages. In The Bacteriophages, R. Calendar, ed., Oxford Press, Chapter 6, 49-65.
  • Choi, K. H., M. C. Morais, D. L. Anderson, and M. G. Rossmann. 2006. Determinants of bacteriophage ø29 head morphology. Structure 14:1723-1727.
  • Morais, M. C., K. H. Choi, J. S. Koti, P. R. Chipman, D. L. Anderson, and M. G. Rossmann. 2005. Conservation of the capsid structure in tailed dsDNA bacteriophages: the pseudoatomic structure of ø29. Mol. Cell 18:149-159.
  • Chemla, Y. R., K. Aathavan, J. Michaelis, S. Grimes, P. J. Jardine, D. L. Anderson, and C. Bustamante. 2005. Mechanism of force generation of a viral DNA packaging motor. Cell 122: 683-692.
  • Anderson, D. and S. Grimes. 2005. The ø29 DNA packaging motor: seeking the mechanism. In Viral Genome Packaging. C. Catalano, ed., Landes Bioscience, Chapter 7.



Institute for Molecular Virology
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