Abstract
HPLC methods are shown to be of predictive value for classification of phytase activity of aggregate microbial communities and pure cultures. Applied in initial screens, they obviate the problems of 'false-positive' detection arising from impurity of substrate and imprecision of methodologies that rely on phytate-specific media. In doing so, they simplify selection of candidates for biotechnological applications. Combined with 16S sequencing and simple bioinformatics, they reveal diversity of the histidine phosphatase class of phytases most commonly exploited for biotechnological use. They reveal contribution of multiple inositol-polyphosphate phosphatase (MINPP) activity to aggregate soil phytase activity, and they identity Acinetobacter spp. as harbouring this prevalent soil phytase activity. Previously, among bacteria MINPP was described exclusively as an activity of gut commensals. HPLC methods have also identified, in a facile manner, a known commercially successful histidine (acid) phosphatase enzyme. The methods described afford opportunity for isolation of phytases for biotechnological use from other environments. They reveal the position of attack on phytate by diverse histidine phosphatases, something that other methods lack.
Highlights
There are four forms of phytic acid which have been identified in nature, myo, neo, scyllo- and D-chiro- that differ in their stereochemical conformation (Figure S1) and association with metal ions as phytates in different soils (Turner et al, 2002)
The undigested phytate and other ‘higher’ inositol phosphates are potent anti-nutrients by virtue of their ability to interfere with protein digestion and to chelate metal ions such as calcium, iron, magnesium, manganese and zinc, reducing their bioavailability
We show how different isolates produce different inositol phosphate profiles from phytate and extend the analysis to soil samples supplemented with phytate to follow the activity of aggregate cohorts of microbes
Summary
There are four forms of phytic acid (inositol hexakisphosphate, InsP 6) which have been identified in nature, myo-, neo-, scyllo- and D-chiro- that differ in their stereochemical conformation (Figure S1) and association with metal ions as phytates in different soils (Turner et al, 2002). Myo-inositol hexakisphosphate (InsP6) garners the most attention from plant scientists It is the principal storage form of phosphorous in plants, seeds and grains representing between 50-85% of the total phosphate in plants and forming as much as 1-5% of the dry weight in many seeds, grains and fruits (Raboy & Dickinson, 1993). Monogastric animals such as swine and poultry are fed diets that are largely cerealand/or grain-based, but they lack sufficient levels of endogenous phytase, a mixed group of phosphatases that dephosphorylate phytate (Pandey et al, 2001). Phytases have become a major sector of a global enzyme market of estimated value ca. $5 billion in 2015, with annual growth estimated at 6-8% from 2016-2020 (Guerrand, 2018)
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