Contacts

Dr Gökhan TolunDr Gökhan Tolun

Email:        gokhan_tolun@uow.edu.au
Phone:      +61 2 4221 3365
Office:       B18.130

Google Scholar

Position
  • Senior Lecturer
Research Interests
  • Studying bio-nano-machines carrying out processes involving nucleic acids, such as DNA recombination, replication, repair, and RNA transcription.
  • My multi-disciplinary research group utilizes
    • Molecular imaging (Electron microscopy)
    • Structural biology (cryo-EM)
    • Biochemistry
    • Molecular Biology

The main technique utilized in my group is electron microscopy (EM). In addition to the state-of-the-art cryo-EM, we also use the classical EM techniques such as shadow-casting (i.e., metal shadowing) and negative staining.

Shadow-casting is a technique ideally suited for visualizing DNA and DNA-protein complexes at the single-molecule level. Below are a few examples: 

DNA Replication

A DNA replication product. Arrows indicate ssDNA gaps along the lagging-strand tail. Inset shows a blow-up of the replication loop visible at the fork.
(Tanner and Tolun et al., EMBO J., 2011)

Electron Micrographs

Electron micrographs of DNA minicircles from the kinetoplast DNA (mitochondrial genome) of TbPIF1-RNAi cells of Trypanosoma brucei. A, relaxed minicircle monomer. B–L, dimers, which differ in number of interlocks. The arrows in I–L mark the small regions where individual double helices are visible. (Liu et al., JBC, 2010)

Mcm2-7

Mcm2-7 complex bound to DNA
(Remus, Beuron, Tolun et al.¸ Cell, 2009)

DNA Recombination

Single strand annealing homologous DNA recombination (SSA) is a process found in virtually all life. It is particularly important in the double-strand DNA (dsDNA) viruses, such as the oncogenic viruses Epstein-Barr Virus (EBV) Kaposi's sarcoma-associated herpesvirus (KSHV), and Herpes Simplex Virus 1 (HSV-1).

SSA is catalysed by a protein complex called a two-component recombinase (TCR), composed of an exonuclease and an annealase. The exonuclease generates a single-strand DNA (ssDNA) overhang, and the annealase binds to this nascent ssDNA and anneals it to a homologous ssDNA strand.

Despite about half a century of research, our understanding of the molecular mechanistic details of SSA is limited. My research group is using a multi-disciplinary approach to better understand how this machinery works, with a focus on cryo-EM.

Single Strand DNA

Simplified schematic representation of single-strand DNA annealing homologous recombination (SSA) catalysed by two-component recombinases. Blue and red lines: DNA, orange circle: nuclease and green spheres: annealase. 

Herpes Simplex Virus

The 3D structure of the Herpes Simplex Virus 1 annealase ICP8 reconstructed from EM images (Tolun et al., NAR, 2013). One of the functional models proposed based on this structure was a winner of the FASEB BioArt Competition 2014. It was featured by: NIH Director’s blog by Francis Collins, NIEHS Environmental Factor, Nature Documentaries, and Los Angeles Times.

We are interested in determining the structures of proteins and protein complexes involved in SSA, using cryo-EM.

DNA Replication

In collaboration with the research groups of Nick Dixon, Antoine van Oijen, and Aaron Oakley, we are studying DNA replication to better understand the molecular mechanistic details of this process.

RNA Transcription

In collaboration with the research group of Mikhail Kashlev (NCI/NIH in MD, USA), we are studying how the transcription machinery assembles and works.

Select Publications
  • Tolun G.*, Vijayasarathy C.*, Huang R, Zeng Y, Li Y, Steven AC, Sieving PA, Heymann JB. Paired octamer rings of retinoschisin suggest a junctional model for cell-cell adhesion in the retina. PNAS, 2016 (2016 May 10;113(19):5287-92. Epub 2016 Apr 25. * Denotes equal contribution).
  • Tolun G., Cheng N., Heymann J.B., Watts N.R., Wingfield P.T., Ludgate L., Hu J., Steven A.C.; Nucleic Acid Content of HBV Capsids Depends on its CTD-Phosphorylation State; Microscopy and Microanalysis, 2013, 19 (S2), 54-55.
  • Hickman, A.B., Ewis, E.H., Li, X., Knapp, J.A., Laver, T., Doss, A.L., Tolun, G, Steven A.C., Grishaev, A., Bax, A., Atkinson, P.W., Craig, N.L., Dyda, F.; “Structural Basis for Transposon End Recognition by Hermes, an Octameric hAT DNA Transposase from Musca domestica”; Cell, 158(2):353-67, Jul 17 2014.
  • Tolun G., Makhov A.M., Ludtke S.J. and Griffith J.D.; “Details of ssDNA annealing revealed by an HSV-1 ICP8-ssDNA binary complex.” Nucleic Acids Res. 2013 Jun;41(11):5927-37.Epub Apr 19 2013.
  • Guo X., Kesimer M.*, Tolun G.*, Zheng X., Dittenhafer-Reed K., Denu J.M., Sheehan J.K., Griffith J.D., Li X.; The NAD+-dependent protein deacetylase activity of SIRT1 is regulated by its oligomeric status; Sci Rep. 2012, 2:640. Epub Sep 7 2012 (* denotes equal contribution).
  • Tanner N.A., Tolun G., Loparo J.J., Jergic S., Griffith J.D., Dixon N.E., and van Oijen A.M.; E. coli DNA replication in the absence of free β clamps; EMBO J., Epub Mar 25 2011.
  • Chowdhury A.R., Wang J., Yildirir G., Liu B., Pappas-Brown V., Tolun G., Griffith J.D., Jensen R.E., Bakshi R., Shapiro T.A., Englund P.T.; The killing of African trypanosomes by ethidium bromide; PLoS Pathog., Dec16;6(12):e1001226; 2010.
  • Liu B., Yildirir G., Wang J., Tolun G., Griffith J.D., and Englund P.T.; TbPIF1, a Trypanosoma brucei mitochondrial DNA helicase, is essential for kinetoplast minicircle replication; J Biol Chem. 5;285(10):7056-66.; 2010 (Epub 2009).
  • Remus D., Beuron F., Tolun G., Griffith J.D., Morris E.P. and Diffley J.F.; Concerted Loading of Mcm2-7 Double Hexamers Around DNA during DNA Replication Origin Licensing; Cell. Nov 13;139(4):719-30. Epub Nov 5 2009.
  • Compton S.A., Tolun G, Kamath-Loeb A.S., Loeb L.A., Griffith J.D..; The Werner syndrome protein binds replication fork and Holliday junction DNAs as an oligomer; J Biol Chem. 283(36):24478-83; Sep 5, Epub Jul 2 2008.
  • Tolun, G. and Myers, R.S.; A real-time DNase assay (ReDA) based on PicoGreen® fluorescence. Nucleic Acids Res., 31, e111; 2003.
  • Feng W., Rodriguez-Menocal L., Tolun G., and D'Urso G.; Schizosacchromyces pombe Dpb2 binds to Origin DNA early in S phase, and is required for chromosomal DNA replication; Mol. Biol. Cell, 14, 3427-3436; 2003.
Abbreviated CV
  • 2017-present Senior Lecturer at the University of Wollongong, NSW, Australia
  • 2012-2017 Postdoctoral Fellow at the National Institutes of Health (NIAMS and NCI), MD, USA
  • 2008-2012 Postdoctoral Fellow at the University of North Carolina, Lineberger Comprehensive Cancer Center, Chapel Hill, NC, USA
Select Awards
  • NIH Fellows Award for Research Excellence (FARE) 2016 competition.
  • FASEB BioArt: Scientific Image & Video Competition 2014.
  • FASEB SRC, Machines on Genes: Nucleic Acid Enzymes 2014, excellent research poster award.
  • NIAMS 2014 Retreat, outstanding poster award.
  • Best talk by a Post-doc or Research Faculty at the UNC Biochemistry and Biophysics departmental retreat, 2011.
  • Mary Poston Award, NC Branch of the ASM, 2010. 
Last reviewed: 24 August, 2018