Abstract
A potent phytase-producing bacterium Bacillus lehensis MLB2 was isolated from bean-grown soil. The optimum conditions recorded after optimization were 24 h incubation time, pH 5.5, 37°C, 2% inoculum level, 0.5% rice bran and 0.5% potassium nitrate. An overall 3.144-fold enhancement in phytase production was achieved after optimization. The use of an inexpensive substrate rice bran and short incubation period make the phytase production cost effective. The purified phytase (152.9 U/mg) had a molecular mass of approximately 98.686 kDa as determined by sodium dodecyl sulphate-polyacryalamide gel electrophoresis and confirmed by liquid chromatography-mass spectrometry, optimum pH of 4.5, and temperature of 37°C. It maintained maximum stability in the acidic region from pH 2.0 to 6.0 and retained 100% at 60°C or below. It showed an enhanced activity in the presence of 5 mM K+ and Na+. Ca2+, Mg2+, and Ba2+ did not have any effect or slightly activate the phytase. Group-specific reagents indicated the presence of cysteine and tryptophan in or near the active site of the enzyme. Better pH and temperature broad range adaptability, strict sodium phytate specificity and low Km value of 0.1232 mM, and in vitro release of a significant amount of orthophosphate from feedstuffs, and thus reduction of environmental phosphorus pollution, make the B. lehensis MLB2 phytase a good candidate for feed additive applicability.
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Abbreviations
- DDT:
-
dithiothreitol
- EDTA:
-
ethylenediaminetetraacetatic acid
- NAI:
-
N-acetyl imidazole
- NBS:
-
N-bromosuccinimide
- NEM:
-
N-ethylmaleimide
- PAGE:
-
polyacryalamide gel electrophoresis
- PMSF:
-
phenylmethylsulphonyl fluoride
- SDS:
-
sodium dodecyl sulphate
- TPTZ:
-
2,4,6-Tris(2-pyridyl)-s-triazine
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More, S.S., Shrinivas, S., Agarwal, A. et al. Purification and characterization of phytase from Bacillus lehensis MLB2. Biologia 70, 294–304 (2015). https://doi.org/10.1515/biolog-2015-0049
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DOI: https://doi.org/10.1515/biolog-2015-0049