• Journal of Microbiology and Biotechnology
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• v.30 no.7
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• pp.1104-1107
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• 2020

The N-terminal domain of the Pseudomonas sp. FB15 phytase increases low-temperature activity and catalytic efficiency. In this study, the 3D structure of the N-terminal domain was predicted and substitutions for the amino acid residues of the region assumed to be the active site were made. The activity of mutants, in which alanine (A) was substituted for the original residue, was investigated at various temperatures and pH values. Significant differences in enzymatic activity were observed only in mutant E263A, suggesting that the amino acid residue at position 263 of the N-terminal domain is important in enzyme activity.

• Journal of Microbiology and Biotechnology
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• v.24 no.10
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• pp.1413-1420
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• 2014

Phytate is an antinutritional factor that impacts the bioavailability of essential minerals such as $Ca^{2+}$, $Mg^{2+}$, $Mn^{2+}$, $Zn^{2+}$, and $Fe^{2+}$ by forming insoluble mineral-phytate salts. These insoluble mineral-phytate salts are hydrolyzed rarely by monogastric animals, because they lack the hydrolyzing phytases and thus excrete the majority of them. The ${\beta}$-propeller phytases (BPPs) hydrolyze these insoluble mineral-phytate salts efficiently. In this study, we cloned a novel BPP gene from a marine Pseudomonas sp. This Pseudomonas BPP gene (PsBPP) had low sequence identity with other known phytases and contained an extra internal repeat domain (residues 24-279) and a typical BPP domain (residues 280-634) at the C-terminus. Structure-based sequence alignment suggested that the N-terminal repeat domain did not possess the active-site residues, whereas the C-terminal BPP domain contained multiple calcium-binding sites, which provide a favorable electrostatic environment for substrate binding and catalytic activity. Thus, we overexpressed the BPP domain from Pseudomonas sp. to potentially hydrolyze insoluble mineral-phytate salts. Purified recombinant PsBPP required $Ca^{2+}$ or $Fe^{2+}$ for phytase activity, indicating that PsBPP hydrolyzes insoluble $Fe^{2+}$-phytate or $Ca^{2+}$-phytate salts. The optimal temperature and pH for the hydrolysis of $Ca^{2+}$-phytate by PsBPP were $50^{\circ}C$ and 6.0, respectively. Biochemical and kinetic studies clearly showed that PsBPP efficiently hydrolyzed $Ca^{2+}$-phytate salts and yielded myo-inositol 2,4,6-trisphosphate and three phosphate groups as final products. Finally, we showed that PsBPP was highly effective for hydrolyzing rice bran with high phytate content. Taken together, our results suggest that PsBPP has great potential in the animal feed industry for reducing phytates.