title: Avian multiple inositol polyphosphate phosphatase is an active phytase that can be engineered to help ameliorate the planet's "phosphate crisis".
abstract: Contemporary phytase research is primarily concerned with ameliorating the problem of inadequate digestion of inositol hexakisphosphate (phytate; InsP6) in monogastric farm animal feed, so as to reduce the pollution that results from the high phosphate content of the manure. In the current study we pursue a new, safe and cost-effective solution. We demonstrate that the rate of hydrolysis of InsP6 by recombinant avian MINPP (0.7 micromol/mg protein/min) defines it as by far the most active phytase found to date in any animal cell (the corresponding activity of recombinant mammalian MINPP is only 0.006 micromol/mg protein/min). Although avian MINPP has less than 20% sequence identity with microbial phytases, we create a homology model of MINPP in which it is predicted that the structure of the phytase active site is well-conserved. This model is validated by site-directed mutagenesis and by use of a substrate analogue, scyllo-InsP6, which we demonstrate is only a weak MINPP substrate. In a model chicken cell line, we overexpressed a mutant form of MINPP that is secretion-competent. This version of the enzyme was actively secreted without affecting either cell viability or the cellular levels of any inositol phosphates. Our studies offer a genetic strategy for greatly improving dietary InsP6 digestion in poultry.
authors.affiliation: Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, NIH, DHSS, Research Triangle Park, PO Box 12233, NC 27709, USA.
authors.forename: Jaiesoon
authors.initials: J
authors.lastname: Cho
authors.forename: Kuicheon
authors.initials: K
authors.lastname: Choi
authors.forename: Thomas
authors.initials: T
authors.lastname: Darden
authors.forename: Paul R
authors.initials: PR
authors.lastname: Reynolds
authors.forename: James N
authors.initials: JN
authors.lastname: Petitte
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Shears
mesh_terms: D010832:6-Phytase / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D002645:Chickens / Q000378:metabolism*; D003247:Conservation of Natural Resources; D008297:Male; D010710:Phosphates / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D010833:Phytic Acid / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*
chemical_list: D010710:Phosphates; D010833:Phytic Acid; D010744:Phosphoric Monoester Hydrolases; D010832:6-Phytase; C064119:multiple inositol-polyphosphate phosphatase