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master's thesis
Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu
Zagreb: University of Zagreb, Faculty of Science, 2009. urn:nbn:hr:217:936343
Marić, Marija
University of Zagreb
Faculty of Science
Department of Biology

Institutional repository: Repository of the Faculty of Science

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Marić, M. (2009). Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu (Master's thesis). Zagreb: University of Zagreb, Faculty of Science. Retrieved from https://urn.nsk.hr/urn:nbn:hr:217:936343

Marić, Marija. "Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu." Master's thesis, University of Zagreb, Faculty of Science, 2009. https://urn.nsk.hr/urn:nbn:hr:217:936343

Marić, Marija. "Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu." Master's thesis, University of Zagreb, Faculty of Science, 2009. https://urn.nsk.hr/urn:nbn:hr:217:936343

Marić, M. (2009). 'Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu', Master's thesis, University of Zagreb, Faculty of Science, accessed 25 November 2024, https://urn.nsk.hr/urn:nbn:hr:217:936343

Marić M. Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu [Master's thesis]. Zagreb: University of Zagreb, Faculty of Science; 2009 [cited 2024 November 25] Available at: https://urn.nsk.hr/urn:nbn:hr:217:936343

M. Marić, "Kloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu", Master's thesis, University of Zagreb, Faculty of Science, Zagreb, 2009. Available at: https://urn.nsk.hr/urn:nbn:hr:217:936343

Metadata
TitleKloniranje i ekspresija gena za SGNH-hidrolazu iz bakterije Streptomyces coelicolor u heterolognom sustavu
Title (english)Cloning and expression of SGNH-hydrolase of Streptomyces coelicolor in heterologous host
AuthorMarija Marić
MentorDušica Vujaklija (mentor)
MentorIvana Ivančić Baće (sumentor)
Committee memberDijana Škorić (predsjednik povjerenstva)
Committee memberDubravka Hranilović (član povjerenstva)
Committee memberNataša Bauer (član povjerenstva)
GranterUniversity of Zagreb
Faculty of Science
(Department of Biology)
Zagreb
Defense date and country2009, Croatia
Scientific / art field,
discipline and subdiscipline		NATURAL SCIENCES
Biology
Abstract
Bakterijske SGNH-hidrolaze su značajna grupa enzima koja još uvijek nije dobro istražena, ni strukturalno ni biokemijski. Izvanstanična lipaza iz obitelji SGNH-hidrolaza iz bakterije Streptomyces coelicolor soja A3(2) (Q93J06, SCO7513) je djelomično biokemijski okarakterizirana. Cilj ovog istraživanja bio je izrada konstrukta gena s dodatkom kodona za 6 histidina na 3' kraju gena te kloniranje i ekspresija gena u bakteriji Streptomyces lividans soja TK23 sa svrhom uspostavljanja učinkovite i brze metode za pročišćavanje proteina. Nakon ekspresije rekombinantnog gena u heterolognom sustavu protein je pročišćen taloženjem s amonijevim sulfatom i afinitetnom kromatografijom na nosaču s imobiliziranim metalom. Prisustvo pročišćenog proteina je dokazano Western analizom, a uočeno je da tijekom pročišćavanja dolazi do gubitka specifične aktivnosti. Iako je prema literaturi ovaj protein označen kao izvanstanični, analiza softverom SignalP 3.0 za predviđanje signalnog slijeda proteina nije prepoznala signalni slijed u SGNH-hidrolazi, što upućuje na mogućnost zadržavanja proteina unutar stanica.
Abstract (english)
Bacterial SGNH-hydrolases are an important group of enzymes which are still not well characterised structurally or biochemically. Extracellular SGNH-hydrolase from Streptomyces coelicolor A3(2) (Q93J06, SCO7513) was earlier partially biochemically described. Aim of this investigation was to construct a gene with an additional codons for 6 histidines on 3' end of the gene. Cloning and expression of the gene Streptomyces lividans TK23 was performed with a purpose of establishing an efficient and quick protein purification method. After the expression of the recombinant gene in heterologous system protein was purified applying ammonium sulfate precipitation and immobilized-metal affinity chromatography. Presence of the purified protein was confirmed with Western analysis. The protein lost specific activity during the purification. Although according to the literature this protein was annotated as extracellular, software SignalP 3.0 failed to recognize a signal sequence in SGNH-hydrolase which implicates a possibility that the protein retains inside the cell.
Keywords
Keywords (english)
Languagecroatian
URN:NBNurn:nbn:hr:217:936343
Study programmeTitle: Molecular Biology
Study programme type: university
Study level: graduate
Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
Type of resourceText
File originBorn digital
Access conditionsOpen access
Terms of useLicence In copyright icon
Created on2018-03-13 11:40:57