Purification and Characterization of an Acidic Polygalacturonase from Grapes and its Potential to Improve Juice Quality
Purification and Characterization of an Acidic Polygalacturonase from Grapes and its Potential to Improve Juice Quality
Zahra Nazir1, Saba Ijaz1, Roquyya Gul2 and Mahjabeen Saleem1,*
ABSTRACT
Pectinolytic enzymes, widespread in plants, fungi and microorganism, gain commercial importance for improving the yield and nutraceutical properties of juice processing industry, degumming of fibre plants and maximum oil recovery. These enzymes break down complex polysaccharide polymers of plant tissues into simpler monomer like D-galacturonic acids. In this study, polygalacturonase from grape skin was purified by salting out with ammonium sulfate, gel filtration on Sephadex G-75 column and ion exchange on Q-Sepharose column chromatography. Polygalacturonase was recognised as a protein with 47kDa molecular weight by SDS-PAGE. The optimum pH of purified polygalacturonase activity was found to be 4.5 and stable within pH range 3.5-5.5. Temperature dependent studies revealed temperature optimum of enzyme to be 40°C and stable up to 60°C. Among substrates, polygalacturonic acid was established as the best substrate for polygalacturonase showing its specificity in the hydrolysis of polysaccharide galacturonate chain. The presence of Na+1 and K+1 enhanced polygalacturonase activity up to 110% and 130%, respectively when used at a concentration of 1mM, however, the enzyme was almost completely inhibited by Pb+2 and Hg+2. Purified polygalacturonase showed pectinolytic effect in juice clarification by attaining maximum clarity (95% transmittance) after incubation at 50°C for 60 min. Total amount of phenolic and free radical scavenging activity of juice was improved after clarification. To understand the effect of juice clarification parameters, response plots based on surface response methodology were developed that showed significant impact of independent variables like enzyme concentration, temperature and incubation time on the dependent variables and improved the overall quality of apple juice. Hence, acidic plant polygalacturonase isolated in this study would be useful as a potential candidate for its biotechnological applications in fruit juice clarification.
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August 2019
Vol. 51, Iss. 4, Pages 1203-1598
