RESUMO
Choline oxidase catalyzes the oxidation of choline to glycine betaine via betaine aldehyde in glycine betaine biosynthesis and betaine acts as an osmolyte. Choline oxidase has attracted a great deal of attention because of its wide application in clinical and its potential use in enzymatic betaine production. Therefore, the development of efficient methods for overexpression of choline oxidase will be very valuable. In the present study, the choline oxidase gene was amplified from a newly isolated Gram-positive soil Arthrobacter globiformis strain HYJE003 and was cloned into a pET expression vector. Furthermore, the culture conditions were optimized for overexpression of cloned choline oxidase gene in different hosts for periplasmic expression of the enzyme. Expression host system Rosetta-gami2(DE3)pLysS yielded more cell-free protein and 20 fold higher active enzyme compared to any other reported studies. Terrific Broth media were found to be yielding the highest cell biomass, by applying the optimized culture conditions and purification strategy 20,902 U of choline oxidase was produced with a specific activity of 95 U/mg. The optimum pH and temperature for the enzyme activity were found to be 7 and 37 °C, respectively. Finally, we have demonstrated efficient bioconversion of betaine using overexpressed and purified choline oxidase enzyme. The enzymatically produced betaine was estimated by the formation of betaine reineckate and we were able to produce 0.83 molar of betaine from one molar of choline chloride. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02960-z.
