@article{Kirillov_Khassenov_Silayev_2017, title={BIOCHEMICAL PROPERTIES OF RECOMBINANT ALKALINE PHOSPHATASE FROM BACILLUS LICHENIFORMIS T5}, url={https://biotechlink.org/index.php/journal/article/view/122}, abstractNote={<p>In this work, recombinant alkaline phosphatase (phoB) from <em>Bacillus licheniformis</em> was successfully expressed in <em>Escherichia coli</em>, purified and biochemically characterized. Gene coding alkaline phosphatase was amplified from genomic DNA of B. licheniformis and cloned into expression vector pET-28c (+). Using the recombinant vector, a BL21 (DE3)/pAlPh strain-producer was obtained with over expression of the gene. Optimal cultivation parameters for producing recombinant alkaline phosphatase have been determined; 10 mg of protein was purified from 1 liter of culture. The activity of the recombinant alkaline phosphatase is 100 U/mg at standard conditions. Biochemical characteristics of recombinant alkaline phosphatase showed that enzyme has maximum activity at pH=10.0 and temperature +60°C. The kinetics of p-nitrophenyl phosphate hydrolysis have been studied, the Michaelis constant Km was 0.91±0.13 mM and the limiting value of the maximal rate of the enzymatic reaction Vmax was 21.4±1.19 mM. Experiments were carried out to determine the dependence of the enzyme activity on various divalent metals.</p>}, number={4}, journal={Eurasian Journal of Applied Biotechnology}, author={Kirillov , S.O. and Khassenov , B.B. and Silayev , D.V.}, year={2017}, month={Dec.} }