Računalne simulacije interakcija dviju domena autotransporterske esteraze EstA iz bakterije Pseudomonas aeruginosa

master's thesis

• Title (english): Molecular dynamics study of functionally relevant interdomain and active site interactions in the autotransporter esterase EstA from Pseudomonas aeruginosa
• Author: Natalia Mrnjavac
• Mentor: Branimir Bertoša (mentor)
• Committee member: Branimir Bertoša (predsjednik povjerenstva)
• Committee member: Ita Gruić Sovulj (član povjerenstva)
• Committee member: Ivana Ivančić Baće (član povjerenstva)
• Committee member: Davor Kovačević (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2015-04-27, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Enzim EstA je funkcionalno GDSL esteraza. Putuje kroz vanjsku membranu bakterije Pseudomonas aeruginosa mehanizmom Va ili autotransporterskim mehanizmom kod kojeg se katalitička domena prenosi u izvanstanični prostor uz pomoć β bačve (autotransporterske domene). U slučaju EstA β bačva i na nju vezana katalitička domena ostaje učvršćena u membrani. Biološki supstrat EstA je nepoznat, ali je poznato da je aktivnost ovog enzima povezana s pokretljivošću bakterijske stanice, stvaranjem biofilmova i produkcijom ramnogalakturonana. Kao u ostalih GDSL hidrolaza, i u aktivnom mjestu EstA se nalazi katalitička trijada i oksioanionska šupljina. U ovom radu su identificirane funkcionalno važne aminokiseline i mreža vodikovih veza u aktivnom mjestu. Uz to, opisane su hidrofobne interakcije i vodikove veze između domena. U odcijepljenoj katalitičkoj domeni je zamijećeno otvaranje aktivnog mjesta kako bi se omogućilo pristajanje tetraedarskog intermedijera, dok je u cjelovitom enzimu EstA s vezanim intermedijerom zamijećena promjena u strukturi heliksa 6 u aktivnom mjestu. Svi rezultati su temeljeni na 100 ns dugim simulacijama molekulske dinamike odcijepljene katalitičke domene i cjelovitog enzima EstA, u oba slučaja sa i bez vezanog tetraedarskog intermedijera.

Abstract (english)

The enzyme EstA is functionally a GDSL esterase. It is transferred through the outer membrane of Pseudomonas aeruginosa by the type Va or autotransporter mechanism, where the transfer of the catalytic domain (the passenger) to the cell exterior is aided by the β barrel domain (the autotransporter). In EstA the barrel remains membrane embedded with the passenger bound to it. The physiological substrate of EstA is unknown, although its activity is known to be related to bacterial cell motility, biofilm formation and rhamnogalacturonan production. As a GDSL hydrolase, the active site of EstA contains a catalytic triad and oxyanion hole. Relevant active site residues, including an active site hydrogen bond network, are described in this work. In addition, interdomain hydrogen bonds and hydrophobic interactions are characterised. Active site opening to fit the tetrahedral intermediate was observed in the isolated passenger domain, while a structural perturbation of the active site helix 6 was noticed when the tetrahedral intermediate was bound in full-length EstA. All results are based on 100 ns long molecular dynamics simulations of the passenger domain of EstA and full-length membrane embedded EstA, both with and without a bound tetrahedral intermediate.

• Keywords: GDSL hidrolaze; katalitička domena; autotransporterska domena; molekulska dinamika
• Keywords (english): GDSL hydrolases; passenger domain; autotransporter domain; molecular dynamics
• Language: croatian
• URN:NBN: urn:nbn:hr:217:602791
• Study programme: Title: Molecular Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-04-24 07:53:18