Karakterizacija substratne specifičnosti in pilov tipa IV pri bakterijah prebavil

Rozman, Vita (author), Vodovnik, Maša (mentor), Accetto, Tomaž (co-mentor)

Pili tipa IV (PT4) so proteinski filamenti na površini nekaterih bakterij in arhej, ki sodelujejo v različnih procesih, med drugim so pri nekaterih bakterijah domnevno pomemben faktor pritrjanja na netopne substrate. V pričujoči študiji smo analizirali sekvence (osnutkov) genomov 118 taksonomsko raznolikih bakterijskih sevov, pretežno izoliranih iz prebavil (kot kontrolo smo vključili tudi nekaj sevov iz zunanjih ekosistemov), z namenom: (1) ugotoviti, ali lahko z »in silico« analizo nabora genov, ki kodirajo encime, aktivne na ogljikovih hidratih (CAZy), pravilno napovemo substratno specifičnost izbranih sevov, (2) ugotoviti, ali obstajajo povezave med nabori CAZy in prisotnostjo ter zgradbo (gruč) genov, ki domnevno kodirajo PT4a. Z algoritmom HMMer smo v osnutkih genomov izbranih sevov identificirali zapise za encime, ki so aktivni na ogljikovih hidratih (CAZy). Na osnovi števila domnevnih monospecifičnih CAZy smo napovedali sposobnost sevov za razgradnjo posameznega substrata. Da se nabor zapisov za CAZy dejansko odraža v substratni specifičnosti sevov, smo nato preverili (in potrdili) še eksperimentalno, z gojenjem 5 izbranih sevov. V drugem delu smo z algoritmoma BLAST in HMMer identificirali gene, ki kodirajo komponente PT4a (gene pil), ter jih glede na arhitekturno ohranjenost in aminokislinska zaporedja ohranjenih genov razvrstili v podskupine. Analiza je pokazala dobro ohranjene genske gruče pil pri mnogih celulolitičnih bakterijah iz različnih habitatov, vendar pa je slednje najti tudi pri nekaterih sevih z drugačnimi napovedanimi specifičnostmi.

• Language: Slovenian
• Keywords: bakterije prebavil, pili tipa IV (PT4), encimi, aktivni na ogljikovih hidratih (CAZy)
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Publisher: [V. Rozman]
• Year: 2017
• PID: 20.500.12556/RUL-95668
• UDC: 579.25:601.4:577.1/.2:601.2:576.34(043.2)
• COBISS.SI-ID: 8817017
• Publication date in RUL: 21.09.2017

Secondary language

• Language: English
• Title: Characterization of substrate specificity and type IV pili of gut bacteria

Abstract

Type IV pili (TFP) are surface filamentous proteins of some bacteria and archea. Among numerous different functions they presumably have an important role in binding to insoluble fiber. In the present study we analyzed (draft) genome sequences of 118 taxonomically diverse bacterial strains, mostly isolated from gut (and some other ecosystems to control for the effect of the environment). The aim of our study was to (1): determine whether »in silico« analysis of genes encoding Carbohydrate Active enZymes (CAZy) reflects in substrate specificity of the selected strains, and (2) determine possible connections among CAZy genes and presence (structure) of putative pil gene clusters. CAZy genes were identified by HMMer algorithm. The number of putative monospecific CAZy was used as an indicator for substrate specificity of the strains. As a “proof of concept”, we verified (and confirmed) that a set of CAZy genes does predict the substrate specificity of 5 selected strains. In the second part of the study pil genes were identified by HMMer and BLAST algorithms. Conserved pil gene clusters were divided into subgroups according to the architecture conservation and amino acid similarities between sequences of pil genes. The results have shown that most of the (confirmed) cellulolytic strains encoded conserved pil gene clusters. However, these were also found in some strains with different predicted specificities.

• Keywords: gut bacteria, type IV pili (TFP), carbohydrate active enzymes (CAZy)