Cloning and expression of SGNH-hydrolase of Streptomyces coelicolor in heterologous host

Marić, Marija (2009) Cloning and expression of SGNH-hydrolase of Streptomyces coelicolor in heterologous host. Diploma thesis, Faculty of Science > Department of Biology.

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Abstract

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.

Item Type: Thesis (Diploma thesis)
Keywords: Streptomyces coelicolor, GDSL/SGNH family, lipase
Supervisor: Vujaklija, Dušica
Co-supervisor: Ivančić-Baće, Ivana
Date: 2009
Number of Pages: 49
Subjects: NATURAL SCIENCES > Biology
Divisions: Faculty of Science > Department of Biology
Depositing User: Silvana Šehić
Date Deposited: 09 Sep 2014 11:20
Last Modified: 09 Sep 2014 11:20
URI: http://digre.pmf.unizg.hr/id/eprint/2675

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