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

Background: Phytases are enzymes capable of degrading phytic acid and used in animal feed supplementation in order to improve digestibility through the release of minerals such as phosphorus. Objective: The main goal of this study was to express and characterize 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 were 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. Conclusion: The results of this article can be considered as innovative and for this reason, they were protected by Intellectual Property Law in Brazil. Take together, the biochemical properties of the rPHY could be useful in future for its industrial application of this enzyme as an additive in the monogastric feed.

Comparative Characteristics of Phytases from Citrobacter freundii and Yersinia intermedia Expressed in Ogataea polymorpha and Pichia pastoris Methylotrophic Yeasts

The genes for bacterial phytases from Citrobacter freundii and Yersinia intermedia were expressed for the first time in a thermotolarant yeast Ogataea polymorpha. A comparative analysis of the properties of recombinant phytases produced by Ogataea polymorpha and Pichia pastoris yeasts was carried out. It was shown that the stability, pH and temperature profiles of the enzyme activities are the same regardless of the host strain. It was proved that O. polymorpha yeast can be used to create producers of feed enzymes and to develop a technology for their cultivation at temperatures above 37 °C. The prospects of using the O. polymorpha yeast for these purposes were evaluated. Ogataea (Hansenula) polymorpha, Pichia pastoris, methylotrophic yeast, thermal tolerance, producer, recombinant phytase The work was financially supported by the Ministry of Science and Higher Education of RF (Project Unique Identifier RFMEFI57917X0145) using the Multipurpose Scientific Installation of All-Russian National Collection of Industrial Microorganisms National Bioresource Center, NRC «Kurchatov Institute»-GosNIIgenetika.

Draft Genome Sequences of Yersinia frederiksenii, Yersinia intermedia, and Yersinia kristensenii Strains from Brazil

ABSTRACT Yersinia enterocolitica-like strains are usually understudied. In this work, we reported the draft genome sequences of two Yersinia frederiksenii, two Yersinia intermedia, and two Yersinia kristensenii strains isolated from humans, animals, food, and the environment in Brazil. These draft genomes will provide better molecular characterizations of these species.