Extraction, purification and determine molecular weight of leucine amino peptidase (LAP) from patients with hypertension

Abstract

The research included isolation and purification of Serum Leucine aminopeptidase from patient with hypertension by gel filtration technique on Sephadex G100 and electrophoresis on SDS-page and determination of the molecular weight of the enzyme by two methods, as well as measurement the kinetic constant [Km,Vmax] , also study the effect of optimum temperature, substrate concentration and pH. The study included 100 patients with hypertension and 75 matched normal individuals were taken as control. The activity of LAP was measured after precipitated by saturation of serum with 45% Ammonium Sulfate concentration, using (Tris-HCl) buffer at pH 8.9. the peak separated from serum by gel filtration technique had apparent molecular weight 32.4 k.Da. SDS electrophoresis techniques had apparent molecular weight of 32.4K.Da. The effect of substrate concentration was found to be increased activity with increasing substrate concentration. The optimum pH was found to be [8.9] and the optimum temperature was 50C°. Michaelis-Menten constant (Km) value for the enzyme was 26.2mM and Vmax value was 8.3IU/ml.