PH AFFECTS MARSH GRAPEFRUIT PECTINESTERASE STABILITY AND CONFORMATION

Thermolabile pectinesterase (TL-PE) and thermostable pectinesterase (T S-PE) of Marsh grapefruit were purified by ion-exchange and affinity c hromatography. The effect of pH on stability and solvent accessible hy drophobicity of both isozymes was studied. TL-PE was inactivated irrev ersibly by 5 min of incubation at pH 2 and pH 12, whereas TS-PE was no t inactivated. Extrinsic 8-anilinonaphthalene-1-sulfonate (ANS) fluore scence intensity was an order of magnitude lower for both PE isozymes than for bovine serum albumin (BSA) at neutral pH. At pH values <3.5, ANS fluorescence of both PE isozymes increased and, at pH 2.0, was hig her than ANS-BSA fluorescence. Fluorescence increase at low pH was gre ater in ANS-TL-PE than in ANS-TS-PE. Activity loss and increase in flu orescence after pH 2 treatment was irreversible for TL-PE, but nearly fully reversible for TS-PE. Resistance to conformational change at low pH by TS-PE probably contributes to its enhanced stability.