Összes szerző

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Sakenov Rauan

az alábbi absztraktok szerzői között szerepel:

Huber Tamás
Comparative Analyses of the Gelsolin Homology Domains of Gelsolin and Flightless-I

Aug 29 - kedd

15:30 – 17:00

I. Poszterszekció

P14

Comparative Analyses of the Gelsolin Homology Domains of Gelsolin and Flightless-I

Tamás Huber^1,3, Péter Gaszler^1,3, Veronika Takács-Kollár¹, Réka Pintér¹, Rauan Sakenov¹, Andrea Teréz Vig¹, Mónika Ágnes Tóth¹, Venukumar Vemula², Marko Ušaj², Alf Månsson² and Beáta Bugyi^1,3

¹University of Pécs, Medical School, Department of Biophysics, Szigeti str. 12, Pécs, H-7624, Hungary

²Linnaeus University, Department of Chemistry and Biomedical Sciences, SE-39182, Kalmar, Sweden

³Regional Committee of The Hungarian Academy of Sciences at Pécs, The Expert Committee of Physics and Astronomy, Spectroscopy Committee

Flightless-I is a unique member of the gelsolin (GSN) superfamily alloying six gelsolin homology (GH) domains and leucine-rich repeats. Flightless-I is an established regulator of the actin cytoskeleton. However, its biochemical activities in actin dynamics regulation are still largely elusive. To better understand its biological functioning, we performed a comparative analysis of GSN and Flightless-I by in vitro fluorescence spectroscopy and single filament TIRF microscopy approaches. We found that Flightless-I can interact with actin and affect actin dynamics in a calcium-independent fashion. Notably, our functional analyses indicate that GSN and Flightless-I respond to calcium differently implying different conformational characteristics of the GH domains in the two proteins. Bioinformatics analyses predict that the sequence elements responsible for calcium activation of GSN are not conserved in the GH domains of Flightless-I. Consistently, the use of intrinsic and extrinsic fluorescent probes revealed that unlike that of GSN the conformational behavior of the GH domains Flightless-I was not significantly affected by calcium-binding. Altogether, our work reveals different calcium-response and predicts distinct modes of activation of GSN and Flightless-I.

Acknowledgment

Activities of Flightless I revealed by acto-myosin based in vitro motility techniques, 2021-4.1.2-NEMZ_KI-2022-00025 (TH); New National Excellence Program of the Ministry for Innovation and Technology ÚNKP-21-3-II-PTE-997 (PG). We thank József Mihály (Institute of Genetics, Biological Research Centre) for the Drosophila Flightless-I plasmids, Tomohito Higashi (Fukushima Medical University) for the human Flightless-I plasmids and Robert C. Robinson (Okoyama University) for the human GSN plasmid.

Szütsné Tóth Mónika Ágnes
Structural features of a disordered protein motif

Aug 29 - kedd

15:30 – 17:00

I. Poszterszekció

P30

Structural features of a disordered protein motif

Mónika Ágnes Tóth¹, Péter Gaszler^{1, 3}, Andrea Vig¹, Veronika Takács-Kollár¹, Illés Csonka², Tamás Huber^{1, 3}, Rauan Sakenov¹, Réka Pintér¹, Beáta Bugyi^{1, 3}

¹University of Pécs, Medical School, Department of Biophysics, Pécs Szigeti str 12. H-7624

²Lajos Nagy High School of the Cistercian Order

³Regional Committee of The Hungarian Academy of Sciences at Pécs, The Expert Committee of Physics and Astronomy, Spectroscopy Committee, Pécs, Hungary

SALS (sarcomere length short) is a Drosophila-specific sarcomeric protein that regulates sarcomere length and organization. Lack of SALS is lethal in embryonic age, possibly due to the shortening of sarcomeric actin filament length or the disruption of their order.

Our bioinformatics analysis suggests that SALS is an intrinsically disordered protein (IDP). IDPs are biologically active proteins, however, do not have a well-defined three-dimensional structure. They possess specific physicochemical properties different from those characteristics for ordered proteins (e.g., amino acid composition, thermal stability, electrophoretic mobility). There is growing attention to studying IDPs for their key roles in diseases or cellular processes. SALS contains two intrinsically disordered protein regions (IDRs), the Wiscott-Aldrich syndrome homology 2 (WH2) domains, composed of a few tens of amino acids. Proteins containing WH2 domains possess actin-binding properties and can exhibit multifunctional character depending on the number and sequence of WH2 domains.

Based on our functional analysis of the SALS WH2 domains (SALS-WH2), both WH2 domains interact with actin and influence actin homeostasis by shifting the monomer:filament ratio towards monomeric actin. The structural properties and conformational dynamics of SALS-WH2 have not yet been described. Therefore, we further aim to characterise these features using in silico and experimental approaches. Our prediction-based results were experimentally verified by fluorescence spectroscopy and thermal analysis.

Acknowledgements

This work was supported by ÚNKP-21-3-II-PTE-997 (PG), University of Pécs, Medical School (MÁT), 2021-4.1.2-NEMZ_KI-2022-00025 (TH). We thank József Mihály (Institute of Genetics, Biological Research Centre, Szeged) for the SALS plasmid.

Takács-Kollár Veronika
Investigation of actin polymerization: methodology and case study

Aug 30 - szerda

15:30 – 17:00

II. Poszterszekció

P31

Investigation of actin polymerization: methodology and case study

Veronika Takács-Kollár¹, Tamás Huber^1,2, Péter Gaszler^1,2, Réka Pintér¹, Rauan Sakenov¹, Andrea Teréz Vig¹, Mónika Ágnes Tóth¹, Beáta Bugyi^1,2

¹ University of Pécs, Medical School, Department of Biophysics

² Regional Committee of The Hungarian Academy of Sciences at Pécs, The Expert Committee of Physics and Astronomy, Spectroscopy Committee, Pécs, Hungary

Actin is one of the most important components of the cytoskeleton that plays a crucial role in many cellular processes. The organization of the actin cytoskeleton is under the control of actin binding proteins (ABPs). Total internal reflection microscopy (TIRFM) is a proper technique for the observation and characterization of actin dynamics from single molecules to a complex system in vitro, it works as a bridge between in vitro and in vivo actin polymerization assays. Furthermore, it is suitable to investigate the role of ABPs in modifying the structural and dynamic properties of actin. Flightless-I (Fli-I) is a member of gelsolin superfamily. It has a unique structure including gelsolin-like domains (GH) and leucine-rich repeats (LRR). Fli-I can interact with actin, however, its biochemical activities in actin dynamics regulation are largely elusive.

The aim of our study is to provide an overview of actin polymerization experiments by fluorescence spectroscopy and TIRFM from basic principles to data/image analysis. We test the applicability of different fluorophores and specify the advantages and limitations of each approach. As a case study, we investigate the effect of wild-type Fli-I and its disease-causing mutations and truncations on actin dynamics. Pyrene-actin-based fluorescence spectroscopy reveals a biphasic effect of Fli-I on actin assembly kinetics, complementary TIRFM assays enlightens the molecular mechanism underlying the activities of Fli-I and its disease-related variants at the single filament level.

Acknowledgment

The research in Hungary was funded by NKFIH within the framework of the project TKP2021-EGA-17 and 2021-4.1.2-NEMZ_KI-2022-00025 ” Activities of Flightless I revealed by acto-myosin based in vitro motility techniques” (TH). We thanks to József Mihály (SZBK, Institute of Genetics, Biological Research Centre) for the Fli-I plasmids.