prikaz prve stranice dokumenta Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus
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master's thesis
Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus
Zagreb: University of Zagreb, Faculty of Food Technology and Biotechnology, 2021. urn:nbn:hr:159:652273
Merkaš, Magdalena
University of Zagreb
Faculty of Food Technology and Biotechnology
Department of Biochemical Engineering
Laboratory for Biochemical Engineering, Industrial Microbiology and Malting and Brewing Technology

Institutional repository: Repository of the Faculty of Food Technology and Biotechnology

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Merkaš, M. (2021). Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus (Master's thesis). Zagreb: University of Zagreb, Faculty of Food Technology and Biotechnology. Retrieved from https://urn.nsk.hr/urn:nbn:hr:159:652273

Merkaš, Magdalena. "Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus." Master's thesis, University of Zagreb, Faculty of Food Technology and Biotechnology, 2021. https://urn.nsk.hr/urn:nbn:hr:159:652273

Merkaš, Magdalena. "Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus." Master's thesis, University of Zagreb, Faculty of Food Technology and Biotechnology, 2021. https://urn.nsk.hr/urn:nbn:hr:159:652273

Merkaš, M. (2021). 'Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus', Master's thesis, University of Zagreb, Faculty of Food Technology and Biotechnology, accessed 11 March 2025, https://urn.nsk.hr/urn:nbn:hr:159:652273

Merkaš M. Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus [Master's thesis]. Zagreb: University of Zagreb, Faculty of Food Technology and Biotechnology; 2021 [cited 2025 March 11] Available at: https://urn.nsk.hr/urn:nbn:hr:159:652273

M. Merkaš, "Synthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus", Master's thesis, University of Zagreb, Faculty of Food Technology and Biotechnology, Zagreb, 2021. Available at: https://urn.nsk.hr/urn:nbn:hr:159:652273

Metadata
TitleSynthesis of mechanistic probes and mutational analysis of the sugar C4/C6 interactions in cytidine 5’-diphosphate-tyvelose 2-epimerase from Thermodesulfatator atlanticus
Title (croatian)Sinteza mehanističkih proba i mutacijska analiza C4/C6 interakcija ugljikohidrata i citidin 5'-difosfat tiveloze 2-epimeraze iz Thermodesulfatator atlanticus
AuthorMagdalena Merkaš
MentorAnita Slavica (mentor)
Committee memberRenata Teparić (član povjerenstva)
GranterUniversity of Zagreb
Faculty of Food Technology and Biotechnology
(Department of Biochemical Engineering)
(Laboratory for Biochemical Engineering, Industrial Microbiology and Malting and Brewing Technology)
Zagreb
Defense date and country2021-07-27, Croatia
Scientific / art field,
discipline and subdiscipline		BIOTECHNICAL SCIENCES
Biotechnology
Abstract
Mechanistic probes for investigation of CDP-Tyvelose 2-Epimerase - like enzyme from Thermodesulfatator atlanticus (TaCPa2E) activity were synthesized. CDP-Lyxose, the mechanistic probe for characterization of C6 interaction with TaCPa2E, was obtained by using chemical approach while CDP-4-Fluoro-4-deoxy-D-Glucose, the mechanistic probe for studying C4 interaction with the enzyme, was yielded by using enzymatic anomeric approach. The later substrate was produced after galactokinase from Streptococcus pneumoniae (GalKSpe4) and UDP-glucose pyrophosporylase from Bifidobacterium longum (UGPase), were produced in E. coli, purified, and therefrom used in the manufacturing of the substrate. Active mutants of TaCPa2E - G206A and D204E were created and used in characterization of the C4/C6 interactions. The highest specific activity towards CDP-Lyxose was determined for wild-type TaCPa2E (0.27 U/ml) and for the two mutants - G206A (0.50 U/ml) and D204E (0.41 U/ml), while CDP-Mannose seems to be better accepted by all three variants (0.02 U/ml for all three forms of the enzyme) than CDP-Glucose. TaCPa2E showed more affinity towards C5 CDP-activated substrate than towards CDP-activated hexoses. Further research might shed more light on CDP-Glucose - and CDP-4-Fluoro-4-deoxy-D-Glucose - enzyme interactions.
Abstract (croatian)
Sintetizirane su mehnističke probe za istraživanje aktivnosti enzima sličnog CDP-Tiveloza 2-Epimerazi iz Thermodesulfatator atlanticus (TaCPa2E). CDP-liksoza, mehanistička proba za karakterizaciju C6 interakcije supstrata s TaCPa2E je dobivena kemijskom sintezom, dok je CDP-4-Fluoro-4-deoksi-D-glukoza, mehanistička proba za istraživanje C4 interakcije supstrata s enzimom, dobivena enzimskom sintezom. Ovaj drugi supstrat je proizveden nakon što su galaktokinaza iz Streptococcus pneumoniae (GalKSpe4) i UDP-glukoza pirofosforilaza iz Bifidobacterium longum (UGPase) sintetizirane u E. coli, pročišćene i onda korištene u proizvodnji ovoga supstrata. Aktivni mutanti TaCPa2E - G206A i D204E također su sintetizirani i korišteni za karakterizaciju C4/C6 interakcija. Najveću specifičnu aktivnost prema CDP-liksozi pokazali su divlji tip TaCPa2E (0.27 U/ml) i dva mutanta - G206A (0.50 U/ml) i D204E (0.41 U/ml), dok je CDP-manoza bolje prihvaćena kao susptrat kod sva tri enzima (0.02 U/ml za sve tri forme ovoga enzima) od CDP-glukoze. Ustanovljeno je da TaCPa2E pokazuje bolju aktivnost prema C5 CDP-aktiviranom supstratu nego prema CDP-aktiviranim heksozama. Potrebna su daljnja istraživanja kako bi se bolje pojasnile interakcije CDP-glukoze i CDP-4-Fluoro-4-deoksi-D-glukoze s ovim enzimom.
Keywords
Keywords (english)
Languageenglish
URN:NBNurn:nbn:hr:159:652273
Study programmeTitle: Molecular Biotechnology
Study programme type: university
Study level: graduate
Academic / professional title: magistar/magistra inženjer/inženjerka molekularne biotehnologije (magistar/magistra inženjer/inženjerka molekularne biotehnologije)
Type of resourceText
File originBorn digital
Access conditionsOpen access
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Created on2021-10-05 11:50:32