Expression and Biochemical Characterization of a Yersinia intermedia Phytase Expressed in Escherichia coli

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Author(s): Mariana Sousa Vieira, Vinícius Valim Pereira, Alice da Cunha Morales Álvares, Lais Moreira Nogueira, William James Nogueira Lima, Paulo Afonso Granjeiro, Daniel Bonoto Gonçalves, Mariana Campos-da-Paz, Sonia Maria de Freitas, Alexsandro Sobreira Galdino*.

Journal Name: Recent Patents on Food, Nutrition & Agriculture

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Background: Phytases are enzymes capable of degrade phytic acid and used in animal feed supplementation in order to impro digestibility throught the release of mineral such phosphorus.

Objective: The main goal of this study was to express and characterize of a Yersinia intermedia phytase expressed in Escherichia coli cells.

Methods: The Y. intermedia phytase gene was synthesized and overexpressed in Escherichia coli cells. The phytase recombinante (rPHY) was purified to homogeneity using a Ni-NTA column. The biochemical and biophysical properties of the rPHY was measured in order to fully characterize the recombinant enzyme. The following patents database were consulted: Espacenet, USPTO, LATIPAT, Patent Scope, WIPO and Google Patents.

Results: The results showed that the rPHY is active at 37-40ºC and presented an optimal pH and temperature of 8.0 and 40 °C, respectively. The phytase rPHY was activated by Cu2+ ion and showed resistance to trypsin and pepsin, retaining 55% of the activity at the ratio of 0.02. Furthermore, the dissociation constant (Kd = 1.1150 ± 0.0087 mM), as estimated by a fluorescence binding assay, suggests a medium affinity of the enzyme with the substrate.

Conclusions: These results of this article can be considerated as innovative and for this reason, them were protected by Intelllectual Property Law in Brazil. Take together, the biochemical properties of the rPHY could be usefull in future for its industrial application of this enzyme as an additive in monogastric feed.

Keywords: phytase, Yersinia intermedia, biochemistry, physical characterization, enzyme kinetics, recombinant enzyme.

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(E-pub Ahead of Print)
DOI: 10.2174/2212798410666181205114153
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