Response Surface Methodology Based Optimization, Partial Purification and Characterization of Alkaline Phosphatase...

The present study was defined to evaluate the effect of a combinational approach of applying phosphate-solubilizing bacteria and alkaline phosphatase for plant growth promotion as a novel strategy. An extracellular phosphatase producing novel Pseudomonas asiatica strain ZKB1 was isolated from ant hill soil. Alkaline phosphatase production was statistically optimized by Plackett-Burman and central composite designs with a yield of 42.45 U/ml and 5.88-fold enhancement. Alkaline phosphatase was purified by column chromatography (DEAE-Cellulose and Sephadex G-100) with 17.55-fold purification and specific activity of 87.77 U/mg. The molecular mass of purified phosphatase was ~ 45 kDa. The optimum pH and temperature were 9.0 and 50 °C, respectively, revealing alkali-thermostability. Phosphatase exhibited the highest specificity toward p-nitrophenyl phosphate disodium salt. Kinetic analysis revealed K_m (0.434 mM) and V_max (264.44 U/mg). Alkaline phosphatase and Pseudomonas asiatica strain ZKB1 as phosphate-solubilizing bacteria were assessed for their ability to induce plant growth in pot experiments with Phaseolus mungo seeds. Seeds soaked in bacterial culture broth and irrigated with increased phosphatase concentration demonstrated better growth with plumule and radical length of 14.8 ± 0.2 cm and 3.5 ± 0.4 cm, respectively. Results were consistent with the combinational approach in terms of enhanced growth. The study suggests the application of alkaline phosphatases in agricultural management, crop improvements, and soil fertility enhancement.