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DOI: 10.5593/SGEM2016/HB63/S08.044

STEPS TOWARDS AN ARTHROBACTER NICOTINOVORANS BASED BIOTECHNOLOGY FOR PRODUCTION OF 6-HIDROXY-NICOTINE

G. Mihalache, M. Stefan, C. Babii, D. Motei, M. Mihasan
Friday 11 November 2016

References: 16th International Multidisciplinary Scientific GeoConference SGEM 2016, SGEM Vienna GREEN Extended Scientific Sessions, www.sgemviennagreen.org, SGEM2016 Conference Proceedings, ISBN 978-619-7105-79-7 / ISSN 1314-2704, 2 - 5 November, 2016, Book 6 Vol. 3, 341-346pp, DOI: 10.5593/SGEM2016/HB63/S08.044

ABSTRACT
As the archetypal agonist of nAchR, nicotine stands up as a powerful scaffold for developing new Alzheimer disease therapeutic agents. In this context, the wide range of nicotine-derivatives produced by Arthrobacter nicotinovorans when grown on nicotine containing media has a huge biotechnological potential. Indeed, the metabolic intermediate 6-Hydroxy-Nicotine (6HNic) produced by A. nicotinovorans pAO1 was shown to bind to nAChRs, and by modulating their function, to sustain spatial memory formation in a rat model of Alzheimer’s disease.
The current work presents the first attempts to produce and isolate 6HNic using a genetically engineered Arthrobacter nicotinovorans strain. The growth and the 6HNic accumulation in a nicotine-containing medium were compared for two strains: 1. A. nicotinovorans pAO1 wild type strain and 2. a genetically engineered A. nicotinovorans pAO1 overexpressing the nicotine-dehydrogenase. The growth curves were fallowed spectrophotometrically and the consumption of nicotine and accumulation of 6HNic in the growth medium was monitored by HPLC.
The growth curve of the genetically engineered strain shows that the bacteria grow slower when compared with the wt. As a result, in the wt strain, the nicotine is quickly depleted from the medium and only low amounts of 6HNic are observed. The overexpression of nicotine-dehydrogenase in the recombinant strain allows for a 5 fold accumulation of 6HNic in the growth medium. Furthermore, the use of inhibitors of the downstream enzymes in the nicotine catabolic pathway leads to a further increase of 6HNiC accumulation.
In conclusion, using the genetically engineered strain for 6HNic production is feasible. Further improvements of the strain are envisioned (i.e. knocking the NDH downstream genes) as well as more data to allow the calculation of the yield and the evaluation of the methods feasibility in an industrial environment.

Keywords: nicotine derivatives, Alzheimer, 6-Hydroxy-Nicotine, Arthrobacter nicotinovorans, biotechnology.

PAPER DOI: 10.5593/SGEM2016/HB63/S08.044, STEPS TOWARDS AN ARTHROBACTER NICOTINOVORANS BASED BIOTECHNOLOGY FOR PRODUCTION OF 6-HIDROXY-NICOTINE

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