KINETICS STUDY OF ALKALINE PHOSPHATASE EXTRACTED FROM KINETICS STUDY OF ALKALINE PHOSPHATASE EXTRACTED FROM Agama agama LIVER IN THE PRESENCE OF MAGNESIUM IONS LIVER IN THE PRESENCE OF MAGNESIUM IONS

Abstract

Alkaline phosphatase (ALP; EC 3.1.3.1) is a metalloenzyme that catalyzes the hydrolysis of phosphate esters under alkaline conditions, playing a vital role in phosphate metabolism, signal transduction, and bone mineralization and is influenced by divalent metal ions such as magnesium (Mg²⁺). It is widely expressed in tissues such as bone, liver, kidney, and intestine, with distinct isoforms including tissue-nonspecific ALP, intestinal ALP, placental ALP, and germ cell ALP and widely used as a biomarker in clinical diagnostics, particularly for liver and bone disorders, due to its tissue-specific isoforms and sensitivity to physiological changes. Kinetics study of alkaline phosphatase (ALP) extracted from Agama agama liver in the presence of magnesium ions investigated the kinetic effects of magnesium ions (Mg2+) on ALP extracted from the liver of Agama agama. ALP activity was measured using para nitrophenyl phosphate (p-NPP) as substrate, monitoring the product para-nitrophenol spectrophotometrically at 405nm. ALP kinetics were assessed at varying p-NPP concentrations in the absence and presence of different Mg2+ concentrations (0.0312, 0.1560, and 0.7800)mg/L. Results showed that in the absence of Mg2+, ALP exhibited moderate activity with a Km of 0.3923mM and Vmax of 0.0237µmol/min. Addition of Mg2+ at low concentration (0.0312mg/L) significantly increased enzyme activity, with Vmax rising to 0.0992µmol/min despite an increased Km of 1.6416mM, indicating enhanced catalytic efficiency but reduced substrate affinity. Higher Mg2+concentrations(0.1560mg/L, 0.7800mg/L) gradually inhibited ALP activity, with decreases in both Vmax and Km, suggesting mixed activation and inhibition effects. The activation constant (Ka) was determined as 1.8359mM. Lineweaver-Burk plots confirmed these kinetic changes. These findings highlight Mg2+ as a critical modulator of ALP activity in reptiles, acting as an activator at low concentrations and an inhibitor at higher levels, consistent with its cofactor role observed in other species, highlighting its dual regulatory role. This study provides insight into reptilian enzyme regulation and contributes to comparative biochemical understanding of metal ion enzyme interactions. Keywords: Alkaline Phosphatase (ALP), Agama agama, Para-nitrophenyl phosphate, Magnesium (Mg2+) ions, UV-Vis spectrophotometer, and Lineweaver-Burk plot.