Seminar Colour Guide:
External Faculty | External Postdoc | Company Representative Science and Society EMBL Distinguished Visitor Lecture Vision2020 Lecture Series Molecular Medicine Seminar | EIPOD Seminar | PSB Seminar | TAC Seminar Hamburg Speaker EMBL-La Sapienza Lecture
|Hamburg Speaker||Abstract: Upon polymerization, actin changes its conformation from|
G-form to F-form (Oda et al, 2009, Nature)(Fig.1), which triggers
(1) A 3.8 Å resolution cryo-EM structure of cofilin-actin
co-filament indicates that cofilin binding brings the actin
molecule into C-form, another conformation. The cofilin-actin
filament structure account for the selective and cooperative
binding of cofilin to ADP-F-actin, formation of cofilin-clusters on
F-actin, severing F-actin at the cluster ends, and accelerated
depolymerization at B- and P-ends.
(2) Crystal structures (at 2.3 Å resolutions) have been
obtained from F2A4 complex (2:4 complex of an actin-binding
protein fragmin (full length) with actin)(Fig.2). Within this
complex, 4 molecules of actin are assembled in the same manner
as in F-actin; two subunits at the B-end are in F-form, two
longitudinal actin-actin contacts as well as one diagonal contact
are canonical. This structure, the first crystal structure of F-form
actin, reveals distinct nature of actin-actin contacts, longitudinal
vs. diagonal contacts.
(3) To our surprise, monomeric F-form actin has been found
in the crystal structure (at 1.15-1.2 Å resolutions) of 1:1 complex
of N-terminal domain-1 of fragmin (F1) with actin (F1A), either
with bound MgAMPPNP (a structural analog of MgATP),
MgADPPi, or MgADP (Fig.3). These structures have allowed us
to investigate plausible reaction paths of actin ATPase by
QM/MM simulations. The reaction path of ATP hydrolysis is very
much similar to those in other ATPase, like myosin. Remarkably,
the main chain configuration is identical between F-ATP and
F-ADP form. However, MD simulations reveal that F-ADP is less
stable, with a pronounced tendency of transition towards G-form.
(4) The monomeric F-form actin is identical to the F-form
actin within F-actin. This means that the G-to-F transition is
induced not only by polymerization but also by a transition
inducer like F1. F1 does not dictate, but selects an actin
conformation. Similarly, we believe that the identical
conformation of F-ATP and F-ADP must be the property of actin itself, not due to F1-binding
hindering in a conformational transition.
Analysis of these recently obtained structures together with all the PDB-deposited actin
structures at atomic resolutions has revealed that actin molecule can take G-, F-, C- (in
cofilin-actin complex) and O-form (in profiling-actin complex) , depending on spatial
relationship between the two rigid bodies , the inner domain (ID) and the outer domain (OD)
excluding the D-loop (Fig.4). Remarkably, MD/PCA analysis reveals that structural
fluctuations of actin monomer are dominated by tendencies of G/F, G/O and F/C transitions,
and that this is shared between monomeric G-form actin and monomeric F-form actin with
bound F1, irrespective of bound nucleotides. (18.1.2018)
|Hamburg Speaker||Abstract: With one-third of mankind infected subclinically, an incidence of nine million new cases of active tuberculosis disease (TB) and two million deaths per year, Mycobacterium tuberculosis, remains the most important bacterial pathogen in the world . This enzyme complex is composed of nine subunits in the stoichiometry of Î±3:Î²3:Î³:Î´:Îµ:a:b:b :c9, and organized in a membrane-embedded FO domain (a:b:b :c9-12) and a water soluble F1 part (Î±3:Î²3:Î³:Î´:Îµ:). Mycobacterial F-ATP synthase has been shown to be essential for growth and survival, which is different from other prokaryotes, where the enzyme is dispensable for growth on fermentable carbon sources and where increased glycolytic flux can compensate for the loss of oxidative phosphorylation . Another special feature of the mycobacterial F-ATP synthase is its inability to establish a significant proton gradient during ATP hydrolysis, and its low or latent ATPase activity in the fast- or slow-growing form [3-4].|
The presentation reveals that at least the three subunits Î±, Î³ and Îµ contribute to the important enzymatic differences of mycobacterial F-ATP synthases in suppression of proton pumping and proton motive force (PMF) formation [4-6], which is essential because dissipating the PMF is lethal to mycobacteria, as well as in ATP formation, employing them as potential drug targets. Our novel atomic structure of the rotating subunit Îµ in solution provides insights into a new mechanism of coupling proton-conduction with ATP synthesis and identifies a second binding site of the TB drug bedaquiline (BDQ, Sirturo®) .
Using a combination of recombineering, ensemble and single molecule assays, solution X-ray scattering, NMR spectroscopy and electron microscopy it will be shown that unique mycobacterial stretches inside the nucleotide-binding subunit Î± and Î³ influence cell growth, ATPase activity and ATP synthesis of the pathogen [4-5].
These data formed a platform to identify and synthesize new compounds, which effectively inhibit ATPase and ATP synthesis of mycrobacterial F-ATP synthase and inhibit growth of Mycobacterium smegmatis, M. bovis as well as M. tuberculosis.
1. World Health Organization (2014) Global Tuberculosis Report 2014, 1-171
2. Cook, G.M., Hards, K., Vilcheze, C., Hartman, T. & Berney, M. (2014) Energetics of Respiration and Oxidative Phosphorylation in Mycobacteria. Microbiol. Spectr. 2, doi:10.1128/microbiolspec.MGM2-0015-2013.
3. Haagsma, A.C., Driessen, N.N., Hahn, M.M., Lill, H. & Bald, D. (2010) ATP synthase in slow- and fast-growing mycobacteria is active in ATP synthesis and blocked in ATP hydrolysis direction. FEMS Microbiol. Lett. 313, 68-74.
4. Hotra, A., Suter, M., BiukoviÄ, G., Ragunathan, P., Kundu, S. Dick, T. & Grüber, G. (2016) Deletion of a unique loop in the mycobacterial F-ATP synthase gamma subunit sheds light on its inhibitory role in ATP hydrolysis-driven H(+) pumping. FEBS J. 283, 1947-1961.
5. Ragunathan, P., Sielaff, H., Sundararaman, L., BiukoviÄ, G., Manimekalai, M.S.S., Singh, D., Kundu, S., Wohland, T., Frasch, W., Dick, T. & Grüber, G. (2017) The uniqueness of subunit Î± of mycobacterial F-ATP synthases: An evolutionary variant for niche adaptation. J. Biol. Chem. 292, 11262-11279.
6. Shin, J., Ragunathan, P., Sundararaman, L., Nartey, W., Kundu, S., Manimekalai, M.S.S., BogdanoviÄ, N., Dick, T. & Grüber, G. (2018) The NMR solution structure of Mycob
|External Faculty Speaker||Abstract: tbd|