PROJECT AARS SUBSTRATE AND EDITING SPECIFICITY

Trajanje / Duration: 1.3.2019. – 31.8.2023.

Financiranje / Funding: Hrvatska zaklada za znanost i Švicarska nacionalna zaklada za znanost / Croatian Science Foundation and Swiss National Science Foundation (a bilateral Croatian-Swiss project)

VODITELJI PROJEKTA / PRINCIPAL INVESTIGATORS: prof. dr. sc. Ita Gruić Sovulj, prof. Nenad Ban

INSTITUCIJE KOJE SUDJELUJU U PROJEKTU / INSTITUTIONS

• Prirodoslovno-matematički fakultet, Sveučilište u Zagrebu / Faculty of Science, University of Zagreb
• ETH Zurich
• Weizmann Institute of Science, Rehovot, Israel

NAZIV PROJEKTA / PROJECT TITLE

Investigation of substrate and editing specificity in tRNA synthetases and the mechanism of antibiotic action

SAŽETAK / ABSTRACT

Proteins are the working horses of the living cell. They are built from twenty amino acids that are attached to one another in a specific order. Aminoacyl-tRNA synthetases (aaRSs) are enzymes that dictate amino acid incorporation into proteins following the information encoded in genes. There are 20 aaRSs in the cell each specific for one amino acid. These enzymes face the difficult task of selecting the designated amino acid from a pool of similar amino acids. Protein functionality strongly depends on the accuracy of amino acid incorporation, making the fidelity of protein synthesis crucial for cell viability.  

This project aims to delineate the evolutionary and mechanistic basis of aaRSs fidelity. AaRS enzymes are very accurate enzymes that double-check that the correct amino acid is chosen. First in the active site of the enzyme and then through additional enzymatic proofreading.  We will investigate which characteristics of aaRSs allow them to be fast and at the same time accurate, and to which degree their enzymatic properties may have evolved to prevent the binding of substrate-mimicking antibiotics into the active sites.

Understanding the molecular mechanisms by which cells ensure high fidelity of protein synthesis under different physiological pressures is of importance for development of new antibiotics and disease treatments.

PROJEKTNI TIM / PROJECT TEAM

• Prirodoslovno-matematički fakultet, Sveučilište u Zagrebu: Ita Gruić Sovulj, Vladimir Zanki, Alojzije Brkić, Aleksandra Maršavelski, Igor Živković, Petra Kozulić, Bartol Božić, Marko Močibob, Morana Dulić
• ETH Zurich: Nenad Ban, Marc Leibundgut, Tanja Schönhut
• Weizmann Institute of Science: Dan Tawfik

PUBLIKACIJE / PUBLICATIONS

1. Brkic A, Leibundgut M, Jablonska J, Zanki V, Car Z, Petrovic Perokovic V, Marsavelski A, Ban N, Gruic-Sovulj I., Antibiotic hyper-resistance in a class I aminoacyl-tRNA synthetase with altered active site signature motif, Nat Commun. (2023)14(1):5498. doi: 10.1038/s41467-023-41244-3. - The publication is selected for the Editors’ Highlights in “Structural biology, biochemistry and biophysics”
2. Zanki V, Bozic B, Mocibob M, Ban N, Gruic-Sovulj I, A pair of isoleucyl-tRNA synthetases in Bacilli fulfills complementary roles to keep fast translation and provide antibiotic resistance, Pro Sci 31 (2022) e4418, doi: 10.1002/pro.4418
3. Zivkovic I, Ivkovic K, Cvetesic N, Marsavelski A, Gruic-Sovulj I., Negative catalysis by the editing domain of class I aminoacyl-tRNA synthetases, Nucleic Acids Res. (2022) 50(7):4029-4041. doi: 10.1093/nar/gkac207.
4. Gruic-Sovulj I, Longo LM, Jabłońska J, Tawfik DS. The evolutionary history of the HUP domain, Crit Rev Biochem Mol Biol. (2022) 57(1):1-15. doi: 10.1080/10409238.2021.1957764. 

KONFERENCIJE / CONFERENCES

1. I Gruić Sovulj, Mupirocin hyper-resistance secured by naturally altered class I signature motif, The IUBMB Focused Meeting on Aminoacyl-tRNA synthetases _ AARS 2023, 4-9.06.2023. Grand Ben, Canada, invited talk
2. I. Živković, A. Maršavelski, I. Gruić Sovulj,  All but one: how negative catalysis shaped evolution of the isoleucyl-tRNA synthetase's editing domain, International conference organized by the Croatian Chemical Society: Solutions in Chemistry, 11-15.11.2022., Sveti Martin na Muri, Croatia - invited talk
3. I Živković, A. Maršavelski, I. Gruić Sovulj, Negative catalysis by isoleucyl-tRNA synthetase: How broad substrate selectivity does not promote futile editing cycles, International Congress of the Croatian Society of Biochemistry and Molecular Biology HDBMB22: FROM SCIENCE TO KNOWLEDGE, 28.09.-1.10.2022., Brela Croatia
4. A. Brkić, B. Božić, M. Leibundgut, N. Ban, I. Gruić Sovulj, Novel insights into the structural basis of antibiotic resistance in type II isoleucyl-tRNA synthetases, 5th Mini Symposium of Section of Medicinal and Pharmaceutical Chemistry, 30.11.2021., Zagreb, Croatia
5. A. Brkić, B. Božić, M. Leibundgut, N. Ban, I. Gruić Sovulj, Structure of a bacterial full­​-length type 2 IleRS reveals the C­​-terminal tRNA-binding domain insertion dispensable for aminoacylation, FEBS Congress: Molecules of Life: Towards new horizons, 3.-8.7.2021. Ljubljana, Slovenia (virtual)
6. I. Gruić Sovulj, Keeping translation canonical: Lessons from aminoacyl-tRNA synthetases, FEBS Congress: Molecules of Life: Towards new horizons, 3.-8.7.2021. Ljubljana, Slovenia (virtual)- Invited talk and charging the session
7. V. Zanki, B. Božić, I. Gruić Sovulj, Isoleucyl-tRNA synthetase carrying antibiotic resistance cannot support sporulation and biofilm formation in Bacillus megaterium, FEBS Congress: Molecules of Life: Towards new horizons, 3.-8.7.2021. Ljubljana, Slovenija (virtualno)
8. A. Brkić, M. Leibundgut, N. Ban, I. Gruić Sovulj, Structure of a bacterial full-length type 2 isoleucyl-tRNA synthetase reveals the C-terminal tRNA-binding domain, Virtual symposium celebrating the 50th anniversary of the Protein Data Bank, 4.-5.5.2021. USA (virtual)
9. A. Brkić, M. Leibundgut, N. Ban, I. Gruić Sovulj, Novel insights into the structural basis of antibiotic resistance in type II isoleucyl-tRNA synthetases, 5. Simpozij studenata doktorskih studija PMF-a, 24.-25.4.2021., Zagreb, Croatia
10. V. Zanki, I. Gruić Sovulj, Izoleucil-tRNA-sintetaza tipa 1 presudna je za sporulaciju i nastanak biofilmova u bakteriji Bacillus megaterium, Simpozij studenata doktorskih studija PMF-a, 28.2.2020., Zagreb, Croatia - the best micro-talk award
11. V. Zanki, I. Gruić Sovulj, Two distinct IleRS proteins in Bacillus megaterium and their possible roles in adaptation to various stress conditions, HDBMB2019, Crossroads in Life Sciences, 25.-28.9.2019., Lovran, Croatia
12. V. Zanki, I. Gruić Sovulj, IleRS2 is responsible for mupirocin resistance of Bacillus megaterium, Bacell 2019, 9.-10.4.2019., Ljubljana, Sloveniaa
13. V. Zanki, I. Gruić Sovulj, Two IleRSs in Bacillus megaterium: is there a difference?, The 3rd COST-sponsored ARBRE-MOBIEU plenary meeting, 18.-20.3.2019., Zagreb, Croatia

OSTALE AKTIVNOSTI / OTHER ACTIVITIES

1. Ita Gruić Sovulj, Isoleucyl-tRNA synthetase: from negative catalysis to antibiotic resistance, Colloquium at the Institute for Biochemistry Leipzig University – virtual event, 15.11.2022.
2. Radni posjet/Working visit. Ita Gruić Sovulj to the Ban lab at ETH Zurich (October 2023.) 
3. Doktorska disertacija/PhD. Thesis. Vladimir Zanki, Različite uloge dviju izoleucil-tRNA-sintetaza iz bakterije Bacillus megaterium u staničnom odgovoru na stres / Different roles of two isoleucyl-tRNA synthetases from Bacillus megaterium in cellular stress response, Faculty of Science University of Zagreb, Zagreb, Croatia, 17.11. 2022.
4. Ita Gruić Sovulj, Isoleucyl-tRNA synthetase: from negative catalysis to antibiotic resistance, Colloquium at the Centre for bacterial cell biology, Newcastle University,  16.05.2022.
5. Radni posjet/Working visit. Vladimir Zanki to the Errington lab at Newcastle University (April-Jule, 2022.) EMBO fellowship 
6. Radni posjet/Working visit. Igor Živković to the Ban lab at ETH Zurich (May-July, 2023.) 
7. Radni posjet/Working visit. Alojzije Brkić to the Ban lab na ETH Zurich (February-April 2021.) 
8. Diplomski rad/Diploma Thesis: Bartol Božić, Kinetička karakterizacija homologno nadeksprimiranih izoleucil-tRNA-sintetaza iz bakterije Bacillus megaterium / Kinetic characterization of homologously overexpressed isoleucyl- tRNA synthetases in Bacillus megaterium,  2020, Faculty of Science University of Zagreb, Zagreb, Croatia, (Mentor: prof. dr. sc. Ita Gruić Sovulj, direct supervisor: Vladimir Zanki, mag. mol. biol.)
9. Radni posjet/Working visit. Alojzije Brkić to the Ban lab na ETH Zurich (September-November 2020.) 
10. Radni posjet/Working visit. Alojzije Brkić to the Ban lab na ETH Zurich (February-March 2020.) 
11. Diplomski rad/Diploma Thesis: Lucija Pavlak, Osjetljivost izoleucil-tRNA-sintetaza iz različitih organizama prema mupirocinu na proteinskoj i staničnoj razini/Mupirocin sensitivity of isoleucyl-tRNA synthetases from various organisms on protein and cellular level, 2019, Faculty of Science University of Zagreb, Zagreb, Croatia(Mentor: doc. dr. sc. Morana Dulić)

NAGRADE / AWARDS

• Vladimir Zanki: Annual Prize for Young Scientists given by the Faculty of Science University of Zagreb (2023)
• Igor Živković: Award Leopold Ruzička given by the Croatian Chemical Society for the extraordinary achievements of young scientists in the field of Chemistry (2022)
• Igor Živković: Annual Prize for Young Scientists given by the Croatian Society for Biochemistry and Molecular Biology (2022)
• Kian Bigović Villi: The University of Zagreb Rector’s Award (2022) (Mentor: prof. dr. sc. Ita Gruić Sovulj, direct supervisor: Vladimir Zanki, mag. biol. mol.)
• Vladimir Zanki: EMBO short-term fellowship
• Bartol Božić: Award Zoran Zgaga given by the Croatian Society of Genetic Engineers for the best diploma thesis in the field of genetic engineering (2020)