Kinetika kaskadne reakcije katalizirane cijelim stanicama mikroorganizma

master's thesis

• Title (english): Kinetics of a cascade reaction catalyzed by whole microbial cells
• Author: Lucija Zovko
• Mentor: Zvjezdana Findrik Blažević (mentor)
• Committee member: Zvjezdana Findrik Blažević (predsjednik povjerenstva)
• Committee member: Marija Vuković Domanovac (član povjerenstva)
• Committee member: Martina Sudar (član povjerenstva)
• Granter: University of Zagreb; Faculty of Chemical Engineering and Technology; Zagreb
• Defense date and country: 2019-09-17, Croatia
• Scientific / art field, discipline and subdiscipline: TECHNICAL SCIENCES; Chemical Engineering

Abstract

Enzimi nude jedan od najboljih načina proizvodnje enantiomerno čistih spojeva zbog svoje izrazite stereoselektivnosti te se iz tog razloga sve više primjenjuju za dobivanje farmaceutika. Kako je dokazano da pogrešan enantiomer može uzrokavati štetne i ozbiljne nuspojave, upravo je kiralnost ključni čimbenik za dobivanje sigurnih i djelotvornih lijekova. U ovome radu proučavana je kinetika kaskadne reakcije katalizirane genetski modificiranim i liofiliziranim cijelim stanicama mikroorganizma Escherichia coli u kojoj se nalaze dva enzima potrebna za provedbu reakcije. Prvi stupanj reakcije kataliziran je enzimom 2-keto-3-deoksi-L-ramonat-aldolazom – YfaU 013. Radi se o reakciji aldolne adicije između natrijevog piruvata i formaldehida te se kao produkt dobiva 4-hidroksi-2-oksobutanoat koji zajedno s L-alaninom sudjeluje u drugom koraku reakcije – reakciji transaminacije (ili reduktivne aminacije) katalizirane enzimom transaminazom – TA 039. Cilj ove kaskadne reakcije je dobivanje aminokiseline L-homoserin koja predstavlja gradivni blok za sintezu drugih važnih i esencijalnih aminokiselina. Korištenjem metode početnih brzina ispitana je kinetika navedene kaskadne reakcije i procijenjene su vrijednosti kinetičkih parametara (Km, Vm). Razvijen je matematički model kaskadne reakcije, te su provedene simulacije pomoću programskog paketa SCIENTIST kako bi se pronašli uvjeti u kojima je moguće dobiti zadovoljavajuću koncentraciju produkta, kao i volumnu produktivnost u reaktoru.

Abstract (english)

Enzymes offer one of the best ways of producing enantiomerically pure compounds due to their great stereoselectivity and therefore they are increasingly being used to obtain pharmaceuticals. Since it is proven that the wrong enantiomer can cause harmful and serious side effects, chirality is a key factor in obtaining safe and effective medications. In this paper, a cascade reaction kinetics has been studied using genetically modified and lyophilized whole cells of Escherichia coli, in which two enzymes that catalyse the reaction were overexpressed. The first step of the reaction was catalysed by the enzyme 2-keto-3-deoxy-L-ramonate-aldolase – YfaU 013. It is an aldol addition between sodium pyruvate and formaldehyde and where 4-hydroxy-2-oxobutanoate is formed which together with L-alanine participates in the second step of reaction called transamination (or reductive amination) catalysed by transaminase – TA 039. The goal of this cascade reaction is to obtain an amino acid L-homoserine which represents a building block for the synthesis of other important and essential amino acids. Kinetics of the cascade reaction were investigated using the initial reaction rate method, and the kinetic parameters (Km, Vm) were estimated from the experimental data. A mathematical model of cascade reaction was developed and simulations were carried out using the program package SCIENTIST to find the conditions at which it is possible to obtain a satisfactory product concentration as well as volume productivity in the reactor.

• Keywords: kinetika; kaskadna reakcija; enzim; L-homoserin; matematički model
• Keywords (english): kinetics; cascade reaction; enzyme; L-homoserine; mathematical model
• Language: croatian
• URN:NBN: urn:nbn:hr:149:225465
• Project: Number: 635595; Title: Sustainable industrial processes based on a C-C bond-forming enzyme platform; Acronym: CARBAZYMES; Leader: Zvjezdana Findrik Blažević; Jurisdiction: eu; Funder: EC; Funding stream: H2020
• Study programme: Title: Applied Chemistry - Graduate study; specializations in: Environmental chemistry and green technologies, Advanced materials and technologies, Applied organic chemistry; Course: Environmental chemistry and green technologies; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra primijenjene kemije (magistar/magistra primijenjene kemije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access; Embargo expiration date: 2021-08-26
• Created on: 2022-02-27 21:48:18