EXPRESSION AND PURIFICATION OF RECOMBINANT PHENYLALANINE 2,3-AMINOMUTASE FROM PANTOEA AGGLOMERANS

Andrea VARGA; Alina  FILIP; László Csaba BENCZE; Péter  SÁTORHELYI; Evelin  SÁNTA-BELL; Beáta G.  VÉRTESSY; László  POPPE; Csaba  PAIZS

EXPRESSION AND PURIFICATION OF RECOMBINANT PHENYLALANINE 2,3-AMINOMUTASE FROM PANTOEA AGGLOMERANS

Authors

Andrea VARGA Faculty of Chemistry and Chemical Engineering, Babeş-Bolyai University, Cluj-Napoca, Romania. Corresponding author: paizs@chem.ubbcluj.ro. https://orcid.org/0000-0002-4290-9226
Alina FILIP Research Center for Enzymology and Applied Biocatalysis, Faculty of Chemistry and Chemical Engineering, Babeş-Bolyai University, Cluj-Napoca, Romania. Emaill: alina.filip@ubbcluj.ro. https://orcid.org/0000-0001-6820-9204
László Csaba BENCZE Biocatalysis and Biotransformations Research Center, Babeş-Bolyai University, Faculty of Chemistry and Chemical Engineering, Cluj-Napoca, Romania. Email: cslbencze@chem.ubbcluj.ro. https://orcid.org/0000-0003-0956-9749
Péter SÁTORHELYI Fermentia Ltd, Budapest, Hungary. Corresponding author: paizs@chem.ubbcluj.ro.
Evelin SÁNTA-BELL Department of Organic Chemistry and Technology, Budapest University of Technology and Economics, Budapest, Hungary. Email: santa-bell.evelin@vbk.bme.hu. https://orcid.org/0000-0002-9083-7757
Beáta G. VÉRTESSY Department of Biotechnology and Food Sciences, Budapest University of Technology and Economics; Institute of Enzymology, Research Centre for Natural Sciences of the Hungarian Academy of Sciences, Budapest, Hungary. Email: vertessy.beata@ttk.mta.hu. https://orcid.org/0000-0002-1288-2982
László POPPE Department of Organic Chemistry and Technology, Faculty of Chemical Technology and Biotechnology, Budapest University of Technology and Economics, Hungary. Email: poppe.laszlo@vbk.bme.hu. https://orcid.org/0000-0002-8358-1378
Csaba PAIZS Faculty of Chemistry and Chemical Engineering, Hungarian Line of Study, Babes-Bolyai University, Cluj Napoca, Romania. Email: csaba.paizs@ubbcluj.ro. https://orcid.org/0000-0002-7403-7098

Keywords:

phenylalanine 2,3-aminomutase, Pantoea agglomerans, optimization, protein expression

Abstract

In the present study, the gene of phenylalanine 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 (18, 25 and 30°C) and the incubation time of PaPAM were examined. The fermentation process was scaled up to 10 L fermentor. Affinity purification conditions were analyzed by SDS-PAGE. The T_m and the activity of the purified enzyme was also investigated.

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STUDIA UBB CHEMIA, Volume 61 (LXI), No. 2, June 2016

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Published

2016-06-30

How to Cite

VARGA, A., FILIP, A. ., BENCZE, L. C., SÁTORHELYI, P. ., SÁNTA-BELL, E. ., VÉRTESSY, B. G. ., … PAIZS, C. . (2016). EXPRESSION AND PURIFICATION OF RECOMBINANT PHENYLALANINE 2,3-AMINOMUTASE FROM PANTOEA AGGLOMERANS. Studia Universitatis Babeș-Bolyai Chemia, 61(2), 7–19. Retrieved from https://studia.reviste.ubbcluj.ro/index.php/chemia/article/view/8304