Identifikacija čimbenika i potencijalnih inhibitora virulencije bakterije Pseudomonas aeruginosa fenotipskom analizom kvasca

master's thesis

• Title (english): Genome-wide identification of Pseudomonas aeruginosa virulence activities and their small molecule inhibitors using phenotypic screens in yeast
• Author: Lana Ljuma
• Mentor: Zoran Zgaga (mentor)
• Granter: University of Zagreb; Faculty of Food Technology and Biotechnology; (Department of Biochemical Engineering); (Laboratory for Biology and Microbial Genetics); Zagreb
• Defense date and country: 2006-06-01, Croatia
• Scientific / art field, discipline and subdiscipline: BIOTECHNICAL SCIENCES; Biotechnology

Abstract

Bakterija Pseudomonas aeruginosa je oportunistički patogen u ljudi, visoko rezistentna na mnoge antimikrobne agense i može razviti visok stupanj višestruke rezistencije na antibiotike, zbog čega se javila potreba za otkrivanjem novih metoda pronalaska ciljnih molekula za djelovanje lijekova. U ovom radu je razvijen novi pristup korištenja kvaščevih stanica kako bi se identificirali i okarakterizirali potencijalni ciljni geni bakterije P. aeruginosa PAO-1. Koristeći GatewayTM tehnologiju, zbirka 505 PAO-1 gena, uključenih u esencijalne stanične procese i virulenciju, bila je prebačena u kvaščev vektor koji je sadržavao inducibilan promotor. Nakon transformacije kvasca Saccharomyces cerevisiae, kvaščeve stanice su bile nacijepljene na podlogu na kojoj je bila inducirana ekspresija PAO-1 gena. Koristeći ovaj pristup, selektirana su četiri bakterijska toksina i šest esencijalnih gena kao potencijalne ciljne molekule za djelovanje lijekova. Geni koji su uzrokovali inhibiciju rasta kvaščevih stanica se trenutno koriste u drugom pretraživanju, kako bi se identificirali kemijski spojevi koji mogu revertirati inhibiciju rasta. Ovaj pristup, temeljen na korištenju cijelih kvaščevih stanica će proširiti primjenu "high through-put" tehnika funkcionalne genomike na patogene mikroorganizme kojima se teško eksperimentalno upravlja.

Abstract (english)

Pseudomonas aeruginosa is an opportunistic human pathogen, highly resistant to multiple antimicrobial agents and can develop high-level multidrug resistance, highlighting the crucial need for new drug target discovery methods. A novel yeast-based approach was developed to identify and characterize potential drug target genes of P. aeruginosa PAO-1. Using the GatewayTM technology, a collection of 505 selected PAO-1 ORFs involved in essential cellular processes and virulence, was transfered to a yeast vector containing an inducible promoter. After transformation in S. cerevisiae, yeast colonies were re-arrayed onto plates where the expression of PAO-1 ORFs was induced. Using such whole-cell approach, four bacterial toxins and six essential genes were selected as potential drug targets. These genes causing growth inhibition are currently being used in a second screen designed to identify chemical compounds capable of reverting the growth inhibition. Such a yeast-based approach will extend the application of high-throughput functional genomics techniques to pathogenic microorganisms that are not readily manipulated experimentally.

• Keywords: ciljne molekule; inhibicija rasta; "high through-put" tehnologija; Pseudomonas aeruginosa; Saccharomyces cerevisiae
• Keywords (english): drug target; growth inhibition; high through-put technology; Pseudomonas aeruginosa; Saccharomyces cerevisiae
• Language: croatian
• URN:NBN: urn:nbn:hr:159:650280
• Study programme: Title: Biotechnology; specializations in: Biochemical-Microbiology; Course: Biochemical-Microbiology; Study programme type: university; Study level: graduate; Academic / professional title: diplomirani inženjer biotehnologije; smjer biokemijsko-mikrobiološki (diplomirani inženjer biotehnologije; smjer biokemijsko-mikrobiološki)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Access restricted to students and staff of home institution
• Created on: 2017-09-07 10:30:10