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Effect of Caffeine and Chlorogenic acid on α–galactosidase Enzyme | ||
| Al-Kunooze Scientific Journal | ||
| Volume 9, Issue 1, June 2023, Pages 15-32 PDF (619.74 K) | ||
| Document Type: Original Article | ||
| Author | ||
| Al-zamely Oda Mizil Yasser, Qasim N. Thewaini, Ameera M.Z. Fadli | ||
| Abstract | ||
| A glycoside hydrolase enzyme known as α-galactosidase (EC 3.2.1.22) hydrolyzes the terminal alpha-galactosyl moieties found in glycolipids and glycoproteins. Bacteria that produce this enzyme are isolated from Erysipelothrix rhusiopathia. Following the release of α-galactosidase from the cells, the purification process involves precipitating the protein with 40% ammonium-sulfate, resuspending it, and dialyzing it using a dialyze tube cut-off number (10–14 KD) overnight against phosphate buffer (pH=7). Finally, the protein is further purified using DEAE-cellulose column chromatography with a (33.4) folds. At 45 °C, the enzyme had maximal activity, while 6.5 °C was its optimum pH. Through the use of SDS-PAGE technique, the molecular weight of the enzyme was estimated to be (55 KD). The enzyme's Km and Vmax were found to be (6.6 mM) and (833.3μmol/min) of protein, respectively, using p-nitrophenyl-α-D-galactopyranoside as the substrate. Following treatments of the purified enzyme with chlorogenic acid and caffeine, studies involving caffeine indicate that the inhibitor has the same affinity for the enzyme as the substrate pNPG. Measurements of the reaction rates at different concentrations of substrate and inhibitor observed a non-competitive inhibition. They showed that the Km value was ineffective (6.66) mM and a decrease in Vmax values (555.5) μmol/min. When we treated chlorogenic acid with α -galactosidase that is purified from Erysipelothrix rhusiopathiae, this substrate acted as uncompetitive inhibition with substrate pNPG as the catalytic site that reflected the enzyme has a single active that results from the change of Km and Vmax (3.22 mM) and (400 μmol/min) respectively. | ||
| Keywords | ||
| coffee; caffeine; chlorogenic acid; α–galactosidase; Erysipelothrix rhusiopathi | ||
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