Alkaline phytase from lily pollen: Investigation of biochemical properties

Abstract

Phytases catalyze the hydrolysis of phytic acid (InsP 6, myo-inositol hexakisphosphate), the most abundant inositol phosphate in cells. In cereal grains and legumes, it constitutes 3–5% of the dry weight of seeds. The inability of humans and monogastric animals such as swine and poultry to absorb complexed InsP 6 has led to nutritional and environmental problems. The efficacy of supplemental phytases to address these issues is well established; thus, there is a need for phytases with a range of biochemical and biophysical properties for numerous applications. An alkaline phytase that shows unique catalytic properties was isolated from plant tissues. In this paper, we report on the biochemical properties of an alkaline phytase from pollen grains of Lilium longiflorum. The enzyme exhibits narrow substrate specificity, it hydrolyzed InsP 6 and para-nitrophenyl phosphate (pNPP). Alkaline phytase followed Michaelis–Menten kinetics with a K m of 81 μM and V max of 217 nmol Pi/min/mg with InsP 6 and a K m of 372 μM and V max of 1272 nmol Pi/min/mg with pNPP. The pH optimum was 8.0 with InsP 6 as the substrate and 7.0 with pNPP. Alkaline phytase was activated by calcium and inactivated by ethylenediaminetetraacetic acid; however, the enzyme retained a low level of activity even in Ca 2+-free medium. Fluoride as well as myo-inositol hexasulfate did not have any inhibitory affect, whereas vanadate inhibited the enzyme. The enzyme was activated by sodium chloride and potassium chloride and inactivated by magnesium chloride; the activation by salts followed the Hofmeister series. The temperature optimum for hydrolysis is 55 °C; the enzyme was stable at 55 °C for about 30 min. The enzyme has unique properties that suggest the potential to be useful as a feed supplement.