Enzymatic characterization of Bacillus subtilis pyruvate kinase

Kovač, Kristina (2012) Enzymatic characterization of Bacillus subtilis pyruvate kinase. Diploma thesis, Faculty of Science > Department of Biology.

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Pyruvate kinase is key enzyme in glycolysis and carbon metabolim. It catalyzes the final step of glycolytic pathway, transferring phosphoryl group of phosphoenolpyruvate (PEP) to ADP, yielding ATP and pyruvate. Reaction is irreversible and it represents important regulatory point in glycolysis. In this research pyruvate kinase is isolated from aerobic Gram-positive bacteria Bacillus subtilis and purified to homogenity. Enzymatic reaction proceeded at pH 7.5 in presence of substrates (ADP and PEP) and ions - K+ and Mg2+. Mg2+ creates a Mg2+-ADP complex with ADP, which acts as a true substrate of the enzyme. Goal of this research was to identify kinetic parameters - Km i Vm of pyruvate kinase. Experimental data given by spectrofotometric method are shown with Michaelis-Menten model of enzyme kinetics. By means of linear plotting, values were calculated to Km=1,2138 mM, Vm=227,062 μmol NADH/min/μg protein for ADP and Km=0,9081 mM, Vm=97,1125 μmol NADH/min/μg protein for PEP. Mentioned results were given by Hanes-Woolf linear plot.

Item Type: Thesis (Diploma thesis)
Keywords: pyruvate kinase, enzymatic characterization, Bacillus subtilis
Supervisor: Franjević, Damjan and Mijaković, Ivan
Date: 2012
Number of Pages: 40
Subjects: NATURAL SCIENCES > Biology
Divisions: Faculty of Science > Department of Biology
Depositing User: Grozdana Sirotic
Date Deposited: 27 May 2014 13:09
Last Modified: 19 Sep 2014 11:02
URI: http://digre.pmf.unizg.hr/id/eprint/1803

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