Stabilization and functional properties of Escherichia coli penicillin G acylase by covalent conjugation of anionic polysaccharide carboxymethylcellulose |
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Authors: | Öztürk Dilek Coskuner Kazan Dilek Erarslan ltan |
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Institution: | (1) The Scientific and Technical Research Council of Turkey, Research Institute for Genetic Engineering and Biotechnology, Marmara Research Center Campus, P.O. Box 21, 41470 Gebze-Kocaeli, Turkey;(2) Department of Chemical Engineering, Marmara University, Göztepe Campus, 81040 Ziverbey–Kadköy, stanbul/, Turkey;(3) Department of Chemistry, Division of Biochemistry, Kocaeli University, 41300 zmit–Kocaeli/, Turkey |
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Abstract: | The stabilization of Escherichia coli penicillin G acylase (PGA) conjugated with carboxymethylcellulose (CMC) against temperature and pH was studied. The 2,3-dialdehyde derivative of CMC obtained by periodate oxidation was covalently conjugated to PGA via Schiff's base formation. The inactivation mechanism of both native and CMC-conjugated PGA appeared to obey first order inactivation kinetics during prolonged incubations at 40–60 °C and in the pH range 4–9. Inactivation rate constants of conjugated enzyme were always lower, and half-life times were always higher than that of native PGA. The activation free energy of inactivation (G
i values) of CMC-conjugated enzyme were found to be always higher than that of native PGA at all temperatures and pH values studied as another indicator of enzyme stabilization. Highest stability of CMC-conjugated enzyme was observed as nearly four-fold at 40 °C and pH 8.0. No changes were observed on the temperature and pH profiles of PGA after CMC conjugation. Lower K
m and higher k
cat values of PGA obtained after CMC conjugation indicates the improved effect of conjugation on the substrate affinity and catalytic performance of the enzyme. |
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Keywords: | Carboxymethylcellulose covalent conjugation enzyme stabilisation Escherichia coli inactivation kinetics penicillin G acylase |
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