Purification and properties of phytate-specific phosphatase from Bacillus subtilis

Abstract

An enzyme which liberates Pi from myo-inositol hexaphosphate (phytic acid) was shown to be present in culture filtrates of Bacillus subtilis. It was purified until it was homogeneous by ultracentrifugation, but it still showed two isozymes on polyacrylamide gel electrophoresis. The enzyme differed from other previously known phytases in its metal requirement and in its specificity for phytate. It had a specific requirement for Ca2' for its activity. The enzyme hydrolyzed only phytate and had no action on other phosphate esters tested. This B. subtilis phytase is the only known phytate-specific phosphatase. The products of hydrolysis of phytate by this enzyme were Pi and myo-inositol monophosphate. The enzyme showed optimum activity at pH 7.5. It was inhibited by Ba2+, Sr2+, Hg2+, Cd2 , and borate. Its activity was unaffected by urea, diisopropylfluorophosphate, arsenate, fluoride, mercaptoethanol, trypsin, papain, and elastase.

Cloning of the thermostable phytase gene (phy) from Bacillus sp. DS11 and its overexpression in Escherichia coli

“…The primary structure of phytase has been reported only for fungi such as A. niger [12^14] and A. fumigatus [15]. However, bacterial phytase genes have not been cloned until now even though two Bacillus phytases have already been puri¢ed and characterized [10,11]. So this is the ¢rst report about the primary structure of bacterial phytase.…”

“…It also occurs in bacteria such as Aerobacter aerogenes [7], Pseudomonas sp. [8], Escherichia coli [9], Bacillus subtilis [10,11]. However, the cloning and sequencing of the phytase gene have been only reported for fungal phytases such as A. niger [12^14], A. fumigatus [15], A. terrus, and Myceliophthora thermophila [16] and there have been no reports about bacterial phytase genes until now.…”

Purification and Properties of a Phytase From Rye

“…The phytases from numerous microorganisms have been investigated. The phytases of Aspergillus jicuum (Hamada 1994;Ullah and Gibson 1987;Ullah 1988), Aspergillus terreus (Yamada et al 1968), Bacillus subtilis (Powar and Jagannathan 1982), Bacillus subtilis (natto) N-77 (Shimizu 1992), Escherichia coli (Greiner et al 1993), Klebsiella Sp. No.…”

Purification and Characterization of a Phytase From Spelt

“…Phytase (myo-inositol hexaphosphate phosp hohydrolase) catalyzes the hydrolysis of phytic acid [myo-inositol( 1,2,3,4,5,6)hexakisphosphate], the major storage form of phosphate in seeds and pollen (Reddy et al 1982), to a series of lower phosphate esters of myo-inositol and phosphoric acid. Two types of phytases are known: 3-phytase (EC 3.1.3.8) and 6-phytase (EC 3.1.3.26), indicating the initial attack on susceptible phosphoester bonds.…”

“…Phytases are present in plants, certain animal tissues and microorganisms (reviewed in Irving 1980;Nayini and Markakas 1986), but only a few phytases have been purified to homogeneity (Yamada et al 1968;Maiti er al. 1974;Powar and Jaganathan 1982;Ullah and Gibson.1987;Segueilha et al 1992;Greiner er al. 1993).…”