Abstract
Herein for the first time a novel acid phosphatase from the seedlings of Cichorium intybus was purified to homogeneity by using various chromatographic techniques (salt precipitation, ion exchange, size exclusion and affinity chromatography) and thermodynamically characterized. The molecular mass of purified enzyme (66 kDa) was determined by SDS-PAGE under denaturing and non-denaturing conditions and by gel-filtration confirmed as dimer of molecular mass 130 kDa. The Michaelis-Menten (K-m) constant for -p-NPP (0.3 mM) and (7.6 mu mol/min/mg) V-max. The enzyme was competitively inhibited by phosphate, molybdate and vanadate. Phenyl phosphate, alpha and beta-glycero-phosphate and-p-NPP were found to be good substrate. When temperature increased from (55 degrees C to 75 degrees C), the deactivation rate constant (k(d)) was increased (0.1 to 4.6 min(-1)) and halflife was decreased from 630 min to 15 min. Various thermal denaturation parameters; change in enthalpy (Delta H degrees), change in entropy (Delta S degrees) and change in free energy (.G degrees) were found 121.93 KJ.mol(-1), 72.45 KJ.mol(-1) and 98.08 KJ.mol(-1) respectively, confirming that acid phosphatase undergoes a significant process of unfolding during deactivation. The biochemical properties of acid phosphatase from C. intybus on the behalf of biological activity and its relationship to pH variations, thermal deactivation and kinetics parameters provide an insight into its novel features. (C) 2020 Elsevier B.V. All rights reserved.