Molekulski mehanizmi bakterijske perzistencije koji se ostvaruju kroz inhibiciju translacije

undergraduate thesis

• Title (english): Molecular mechanisms of bacterial persistence mediated by translation inhibition
• Author: Gracia Štefanić
• Mentor: Ita Gruić-Sovulj (mentor)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2020, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Cilj ovog rada jest objasniti bakterijsku perzistenciju, sustav toksin-antitoksin i načine njegovog djelovanja, proces translacije i načine njene inhibicije te postizanje perzistencije toksinom HipA i model njegova djelovanja. Perzistencija je stanje usporenog rasta bakterije u kojem ona postaje tolerantna na antibiotike i druge okolišne stresne uvjete. Pokazano je da perzistencija olakšava nastanak rezistencije i glavni je uzrok kroničnih infekcija te u svrhu razvitka ciljane terapije važno je razjasniti njen mehanizam. Genski krivac za bakterijsku perzistenciju jest sustav toksin-antitoksin. Aktivacijom toksina inhibira se najčešće proces translacije te stanica ulazi u dormantno stanje usporenog rasta i postaje perzistentna. Inaktivacijom toksina stanica može ponovno uspostaviti normalan rast, ali u tom slučaju gubi perzistenciju. Prvi otkriveni gen perzistencije u E. coli je hipA koji kodira za serin/treonin kinazu i inhibira translaciju, transkripciju i replikaciju. Dio je hipBA operona koji još čini gen za antitoksin HipB. Po početnom modelu mehanizma djelovanja HipA, predloženo je da on fosforilira elongacijski faktor EF-Tu, koji dovodi aminoacilirane tRNA na ribosom, i time inhibira translaciju. Međutim, eksperimentom in vitro u kojem je korišten γ[32P]ATP pokazano je da HipA ne fosforilira EF-Tu. Daljnjim eksperimentima opisanim u radu utvrđeno je da HipA fosforilira glutamil-tRNA-sintetazu i time dolazi do inhibicije translacije i aktivacije enzima RelA koji stvara (p)ppGpp. Pokazano je da fosforilacija ovisi o konformaciji glutamil-tRNA-sintetaze. S obzirom na to da HipA uvijek fosforilira na konzerviranom Ser239 koji je dio fleksibilne omče, vezanjem tRNAGlu mijenja se konformacija i Ser239 postane više izložen i dostupan HipA.

Abstract (english)

Aim of this review is to explain bacterial persistence, toxin-antitoxin module and the way it works, protein translation and possible mechanisms of its inhibition and molecular mechanisms of bacterial persistence mediated by HipA. Baterial persistence is a dormant state in which bacteria tolerates many antibiotics and other stressful conditions. It is known that bacterial persistence promotes evolution of antibiotic resistence and it is the main cause of many chronic infections. In order to develop targeted therapy it is important to understand molecular mechanism by which bacteria become persistant. Genes which cause persistance are toxin-antitoxin modules. Toxin activation usually inhibits translation after which cell enters physiological state of slow growth and becomes persistant. If the toxin is inactivated then the cell restores its normal growth but is no longer persistant. First gene discovered in E. coli that causes persistance is hipA. It codes for serine/threonine kinase which inhibits translation, transcription and replication. Together with hipB (gene for anitoxin) is part od hipBA operon. The first model of persistence mediated by HipA suggested that HipA inhibits translation by phosphorylating EF-Tu, the elongation factor which brings charged tRNA molecules to the ribosome. However, in vitro experiments in which they used γ[32P]ATP results did not confirm the initial model. The further experimental work revealed that HipA phosphorylates glutamyl-tRNA-synthetase (GltX) which results in translation inhibition and activation of the enzyme RelA which produces (p)ppGpp. Finally, the results showed that phosphorylation depends on conformation of GltX. HipA always phosphorylates on Ser239 which is part of a conserved flexible loop that changes conformation when tRNAGlu binds and Ser239 becomes more exposed.

• Keywords: bakterijska perzistencija
• Language: croatian
• URN:NBN: urn:nbn:hr:217:215179
• Study programme: Title: Molecular Biology; Study programme type: university; Study level: undergraduate; Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) molekularne biologije (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2020-09-25 10:21:24