Kinetics and thermodynamic stability of native and chemically modified acid invertase: Extracted from yellow pea (Lathyrus aphaca) Seedlings

Abstract

In this work, an acid invertase isolated from Lathyrus aphaca seedlings is purified, and its thermal properties are investigated. Acid invertase was purified through salt fractionation, CM-cellulose, and Ultrogel ACA-44 chromatography. Using SDS-PAGE, the consistency of the isolated enzyme was verified. The enzyme weighs 29 kDa at the molecular level. Acid invertase functions best at a pH of 3.0 and a temperature of 45 °C. Activation energy of 29.5 kJ mol−1 and Km and Vmax values of 0.5 mM and 119.7 µmol. min−1. mg−1 of protein was found for the isolated enzyme. It was shown that the mercaptide-forming agent p-chloromercuribenzoic acid (PCMB; 0.5 mM) and the enzyme's activity were both modestly increased by Ca2+, Mn2+, and Mg2+ ions but inhibited by Hg2+, Cd2+, and Pb2+ ions. At elevated temperatures, the invertase exhibited an increase in thermostability due to an increase in activation entropies (∆Sº) and a decrease in activation enthalpies (∆Hº). The interactions between 4 M urea and α-chymotrypsin were tetraphasic, causing periodic increases and decreases in invertase activity. It also suggests a potential explanation for acid-invertase thermal inactivation at high temperatures. This enzyme might be a catabolite-resistant invertase to produce high-gravity ethanol or fructose syrup.