Anna Roujeinikova's laboratory

Anna Roujeinikova Associate Professor Phone: +61 3 9902 9194 Fax: +61 3 9902 9222 E-mail: Anna.Roujeinikova@med.monash.edu.au Postal address: Building 76, School of Biomedical Sciences, Monash University, Clayton, VIC 3800, Australia

BSc, Natural Sciences (1994, Moscow Institute of Physics and Technology, Russia)
MSc, Applied Physics and Mathematics (1997, Moscow Institute of Physics and Technology)
PhD, Structural Biology (2002, University of Sheffield, UK)
Wellcome Trust Research Career Development Fellowship (2006-2009, University of Manchester, UK)

Keywords: X-ray crystallography, protein structure, molecular machines, protein-protein and protein-substrate recognition, bacterial motility, bacterial flagellar motor, virulence factors.

The projects in the lab use a structural biology approach to gain molecular understanding of how medically-important bacteria, such as Helicobacter pylori and Neisseria gonorrhoeae cause disease.

Understanding the function of the motility protein B component of the bacterial flagellar motor (BFM)

The bacterial flagellar motor is a membrane-embedded molecular machine that rotates filaments, providing a propulsive force for bacteria to swim toward nutrients, optimal temperatures, or other factors that favour survival. Motility by flagellar motor is essential for the survival, chemotaxis and virulence of many pathogenic bacteria. The chosen model system, the carcinogenic bacterium Helicobacter pylori, uses the flagellar motor to drill into the mucus layer of the stomach and move towards the epithelial surface, where it colonizes. The molecular mechanism of torque (turning force) generation is being investigated through the study of the properties and three-dimensional structure of the individual components of the motor's stator unit. The highly dynamic nature of the stator complex creates the biggest challenge in obtaining its structural information. We are taking both top-down and bottom-up approaches to this problem, combining data from the molecular genetics studies, cross-linking, X-ray protein crystallography, electron microscopy, peptide amide hydrogen/deuterium exchange coupled with liquid chromatography and mass spectrometry. We have recently determined the first crystal structure of the protein domain that anchors the proton-motive-force-generating mechanism of the bacterial flagellar motor to the cell wall, and formulated a model of how the stator attaches to peptidoglycan. The ongoing research aims to unravel the mechanism of stator assembly, anchoring to the peptidoglycan and force generation.

Electron density for N-acetylmuramic acid bound to the MotB peptidoglycan-binding domain and the model for the glycan chain binding.

Structural biology of major virulence factors of the carcinogenic bacterium Helicobacter pylori (in collaboration with Prof Trevor Lithgow, Dr Terry Kwok)

Pathogenic strains of Helicobacter pylori, associated with the development of adenocarcinoma in humans, inject CagA protein into gastric epithelial cells, where it interacts with many different host cell proteins, interfering with signalling pathways that regulate the cell growth and motility. Detailed characterization of the CagA structure and interactions will be undertaken for elucidation of its role in gastric carcinogenesis. In complementing experiments, the in-vivo activity of CagA fragments (e.g. effect on cell morphology and motility, interaction with various partner molecules) will be assessed. We are also interested in investigating the structure/function relationships in VacA – a multifunctional H. pylori toxin that induces a variety of cytopathic effects in mammalian cells. We aim to understand, at the molecular and structural level, how VacA is secreted and activated, how it inserts into host membranes to form channels and how it interacts with its receptors and the cytosolic proteins.

Understanding bacterial stress response mechanisms (in collaboration with Prof John Davies)

Interaction between heat shock sigma factor RpoH (sigma32) and molecular chaperone DnaK is strongly implicated in the stress response and virulence in N. gonorrhoea. We will apply a range of structural biology techniques to understand the molecular mechanisms that underpin the specific RpoH-DnaK recognition and stress-induced complex dissociation. We are also interested in investigating the potential of DnaK as a drug target. Selective, peptide-based, antibacterial inhibitors of DnaK may be able to kill some of the important pathogenic bacteria by stalling their essential protein repair machinery.

Pyrrhocoricins act competitive inhibitors of E. coli DnaK

Joining the lab:

Prospective PhD students:

A PhD scholarship is open to Australian citizens, New Zealand citizens, and Australian permanent residents who hold an Australian or New Zealand bachelors degree with first class honours or overseas qualifications deemed equivalent by Monash university (please enquire if you are not sure). Applications from candidates currently completing an honours degree are also welcome. Applicants must meet English language proficiency standards. Enquiries and applications can be sent to Anna.Roujeinikova@med.monash.edu.au

A range of other Ph.D. scholarships are open to Australian and international students; specific information on how to apply to the Monash postgraduate programme, eligibility and application deadlines can be found here. International students can also apply through the CCV, Endeavour, CSIRO or ESF schemes.

Prospective postdoctoral fellows:

Australian postdoctoral researchers and PhD students in their final year are encouraged to apply through the available fellowship schemes (ARC Australian postdoctoral fellowship, NHMRC training (postdoctoral) fellowship). International candidates should consult HFSP and EMBO web pages for open fellowship schemes. G08 European Fellowships are available to candidates from EU. US citizens are invited to apply through the NSF International Research Fellowships Programme, AAA or LSRF scheme

Recent publications

Liebscher, M. and Roujeinikova, A. (2009) Allosteric coupling between the lid and the interdomain linker in DnaK revealed by inhibitor binding studies. J. Bacteriol. 191, 1456-1462.

Toogood, H.S., Hare, V., Fryszkowska, A., Fisher, K., Roujeinikova, A., Heyes, D.J., Leys, D., Stephens, G.M., Gardiner, J. and Scrutton, N.S. (2008) Structure-based insight into the asymmetric bioreduction of the C═C double bond of α,β-unsaturated nitroalkenes by pentaerythritol tetranitrate reductase. Adv. Synth. Catal. 350, 2789-2803.

Hothi, P., Hay, S., Roujeinikova, A., Sutcliffe, M.J., Lee, M.., Cullis, P.M. and Scrutton, N.S. (2008) Driving force analysis of proton tunnelling across a reactivity series for an enzyme-substrate complex. ChemBioChem. 9, 2839-2845.

Roujeinikova, A. (2008) Cloning, purification and preliminary X-ray analysis of the C-terminal domain of Helicobacter pylori MotB. Acta Cryst. F 64, 277-280.

Hothi, P., Roujeinikova, A., Sutcliffe, M.J., Cullis, P., Leys, D. and Scrutton, N.S. (2007) Isotope effects reveal that p-substituted benzylamines are poor reactivity probes of quinoprotein mechanism for aromatic amine dehydrogenase. Biochemistry 46, 9250-9259.

Roujeinikova, A., Hothi, P., Scrutton, N.S. and Leys, D. (2007) New insights into the reductive half-reaction mechanism of aromatic amine dehydrogenase revealed by reaction with carbinolamine substrates. J. Biol. Chem. 282, 23766-23777.

Roujeinikova, A., Simon, J.W., Gilroy, J., Stuitje, A.R., Rice, D.W., Slabas, A.R. and Rafferty J.B. (2007) Structural studies of fatty acyl-(acyl carrier protein) thioesters reveal a hydrophobic binding cavity that can expand to fit longer substrates. J. Mol. Biol. 365, 135-145.