Expression and purification of recombinant phenylalanine 2,3-aminomutase from Pantoea agglomerans

Varga, Andrea and Filip, Alina and Bencze, László Csaba and Sátorhelyi, Péter and Bell, Evelin and Vértessy G., Beáta and Poppe, László (2016) Expression and purification of recombinant phenylalanine 2,3-aminomutase from Pantoea agglomerans. STUDIA UNIVERSITATIS BABES-BOLYAI CHEMIA, 61 (2). pp. 7-19. ISSN 1224-7154

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Abstract

In the present study the gene of phenylalamine 2,3-aminomutase from Pantoea agglomerans (PaPAM) was cloned into pET-19b vector and used for its expression in competent Escherichia coli cells. The recombinant plasmid, PaPAM-pET-19b, was transformed into competent E. coli strain BL21(DE3)pLysS cells. Overnight culture of the transformed bacteria was induced by the addition of isopropylthio-β-D-galactoside (IPTG) to the final concentrations of 0.1, 0.5 and 1 mM. Also, the effects of different temperatures (15, 20, 25 and 30°C) and the incubation time of PaPAM. The fermentation process was scaled up to 10 L fermentor. Affinity purification conditions were analyzed by SDS-PAGE. The Tm and the activity of the purified enzyme was also investigated.

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