Összes szerző

Nyitrai Miklos

az alábbi absztraktok szerzői között szerepel:

Barkó Szilvia
Direct binding of fluorescent vancomycin to MreB

Aug 29 - kedd

15:30 – 17:00

I. Poszterszekció

P03

Direct binding of fluorescent vancomycin to MreB

Beáta Longauer¹, Emőke Bódis¹, Zoltán Gazdag⁴, Miklós Nyitrai^1,2,3 and Szilvia Barkó^1,2,3

¹ University of Pécs, Medical School, Department of Biophysics

² MTA-PTE Nuclear-Mitochondrial Interactions Research Group

³ University of Pécs, Szentágothai Research Center

⁴ University of Pécs, Faculty of Sciences, Department of Molecular Biology and Microbiology

The discovery of antibiotics is one of the greatest discoveries in human history. It is also known that bacteria have developed a serious arsenal of weapons to resist antibiotics. As a result, despite the availability of many antibiotics with very different mechanisms of action, the number of antibiotic-resistant bacterial species is increasing. The result is a worldwide crisis which mankind currently seems powerless to tackle.

In our study we describe a novel potential target in bacteria, which is essential for bacterial survival. MreB, which has major role in organising cell wall synthesis and is found in every bacterium, can be inactivated with a MreB-specific drug A22. This target, although it affects a component of the cell wall, is fundamentally different from the antibiotics used so far.

In our studies, we confirmed the binding of Bodipy-vancomycin to the MreB protein by steady-state anisotropy measurements, which showed an affinity on the order of micromolar. The anisotropy further increased upon exposure to native vancomycin, confirming the previous observation that vancomycin molecules can form supercomplexes with each other and with target proteins. Our light-scattering measurements suggest that vancomycin, like other proteins, can induce aggregation of MreB, but this effect is not observed for ATP-bound protein. This supports our previous observations that nucleotide binding plays a crucial role in MreB stabilisation. Our microbiological and fluorescence microscopy results show that A22 promotes the uptake of vancomycin into cells, which may explain why a synergistic effect between A22 and vancomycin in inhibiting E. coli cell division is observed.

All these observations suggest that antimicrobials based on the mechanism of action of A22 could play a key role in the future in the fight against resistant bacterial strains.

Lukács András
Conformational flexibility in a photoactivated adenylate cyclase studied by fluorescence spectroscopy and solution X-ray scattering

Aug 29 - kedd

09:10 – 09:30

Molekuláris biofizika

E03

Conformational flexibility in a photoactivated adenylate cyclase studied by fluorescence spectroscopy and solution X-ray scattering

1,2, , , András Kengyel, Matteo Levantino, Caroline Maas, Dihia Moussaoui, Ildikó Pécsi, Petra Pernot, Kevin Pounot, Giorgio Schiro, Mark Tully, Kinga Ujfalusi-Pozsonyi, Jovana Vitas, Martin Weik, András Lukács

Sofia Maria Kapetanaki^1,2, Emőke Bódis¹, Miklós Nyitrai¹, András Kengyel¹, Matteo Levantino³, Caroline Maas², Dihia Moussaoui³, Ildikó Pécsi¹, Petra Pernot³, Kevin Pounot^2,3, Giorgio Schirò², Mark Tully³, Kinga Ujfalusi-Pozsonyi¹, Jovana Vitas², Martin Weik², András Lukács¹

¹ Department of Biophysics, Medical School, University of Pécs, 7624 Pécs, Hungary

² CEA–Institut de Biologie Structurale, Grenoble, 38044 France

³ European Synchrotron Radiation Facility, Grenoble, 38043 France

The photoactivated adenylate cyclase from the photosynthetic cyanobacterium Oscillatoria acuminata OaPAC is a homodimeric enzyme comprising of a N-terminal domain that senses blue light using flavin (BLUF) [1] and a C-terminal class III adenylate cyclase (AC) domain that catalyses the formation of cAMP from ATP (adenosine triphosphate) [2,3]. cAMP is a universal regulator of metabolism and gene expression in all life forms [4]. Modulating the cellular concentration of cAMP has emerged in the focus of modern optogenetic applications and therapeutic approaches. Recent crystallographic studies have indicated that the activation mechanism of OaPAC involves only small movements. In this study [5], we apply small-angle X-ray scattering (SAXS) [6] and time-resolved solution X-ray scattering [7] in combination with other biophysical techniques to investigate the substrate induced-conformational changes of OaPAC in solution. The implications of our work to the function of the enzyme are discussed.

Acknowledgment

A.L. acknowledges funding from the Hungarian National Research and Innovation Office (K-137557) and was supported by PTE ÁOK-KA-2021

References

[1] Fujisawa, T. and Masuda, S. (2018) Light-induced chromophore and protein responses and mechanical signal transduction of BLUF proteins Biophys. Rev. 10, 327-337.

[2] Ohki, M. et al. (2016) Structural insight into photoactivation of an adenylate cyclase from a photosynthetic cyanobacterium Proc. Natl. Acad. Sci. 113, 6659-6664.

[3] Ohki, M. et al. (2017) Molecular mechanism of photoactivation of a light-regulated adenylate cyclase Proc. Natl. Acad. Sci. 114, 8562-8567.

[4] Zaccolo, M., Zerio, A., and Lobo, M.J. (2021) Subcellular Organization of the cAMP Signaling Pathway Pharmacol.Rev. 73, 278–309.

[5] Kapetanaki, S.M. et al. (unpublished results)

[6] Da Vela Stefano and Svergun, D. (2020) Methods, development and applications of small-angle X-ray scattering to characterize biological macromolecules in solution Curr. Res. Struct. Biol. 2, 164-170.

[7] Cho, H.S., Schotte, F., Stadnytskyi, V., and Anfinrud, P. (2021) Time-resolved X-ray scattering studies of proteins Curr. Opin. Struct. Biol. 70, 99-107.