Molecular and Biochemical Characteristics of β-Propeller Phytase from Marine Pseudomonas sp. BS10-3 and Its Potential Application for Animal Feed Additives
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Phytate is an antinutritional factor that impacts the bioavailability of essential minerals such as Ca2+, Mg2+, Mn2+, Zn2+, and Fe2+ 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 β-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. Structurebased 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 Ca2+ or Fe2+ for phytase activity, indicating that PsBPP hydrolyzes insoluble Fe2+-phytate or Ca2+-phytate salts. The optimal temperature and pH for the hydrolysis of Ca2+-phytate by PsBPP were 50°C and 6.0, respectively. Biochemical and kinetic studies clearly showed that PsBPP efficiently hydrolyzed Ca2+-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.
Abstract
Phytate is an antinutritional factor that impacts the bioavailability of essential minerals such as Ca2+, Mg2+, Mn2+, Zn2+, and Fe2+ 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 β-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. Structurebased 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 Ca2+ or Fe2+ for phytase activity, indicating that PsBPP hydrolyzes insoluble Fe2+-phytate or Ca2+-phytate salts. The optimal temperature and pH for the hydrolysis of Ca2+-phytate by PsBPP were 50°C and 6.0, respectively. Biochemical and kinetic studies clearly showed that PsBPP efficiently hydrolyzed Ca2+-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.
참고문헌 (18)
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