A CONTRIBUTION TO THE CHARACTERIZATION OF PROTOPECTINASE SE, AN ENDOPOLYGALACTURONASE WITH PECTIN-RELEASING ACTIVITY FROM Geotrichum klebahnii
Hydrolysis of polygalacturonic acid (1.8 g.L−1 ) with 1.25 mg.L−1 of a commercial preparation (Pectinase SE from Shikibo Ltd., Japan) containing protopectinase SE (PPase-SE) activity for 80 min released a reducing power equivalent to ≈ 400 mg.L−1 of galacturonic acid monohydrate, yielding oligogalacturonates with an average polymerization degree of around 4. Thermostability of PPase-SE was positively affected by enzyme concentration. Enzyme activity of a solution containing 12.5 mg.L−1 of Pectinase SE in 20 mM sodium acetate buffer, pH 5.0, quickly dropped to less than 20 % after 80 s of vortexing. Enzyme pre-incubation (37°C, 30 min) with Ca2+, Mg2+, Co2+, Cu2+, Fe2+, Zn2+ and Mn2+ (0.1, 1.0 and 10.0 mM) did not affect the activity except in the case of Cu2+ (10 mM). Ca2+ (2.5 mM) and Mg2+ (2.5 mM) in the reaction mixture caused inhibition and activation, respectively. Ca2+ inhibition was partially reverted by Mg2+. The enzyme is not inhibited by products. Km and Vmax value for PGA was determined to be 0.198 ± 0.013 g.L−1 and 0.0509 ± 0.0032 µmol.mL−1 .min−1 respectively. This Km value is one of the lowest reported for microbial PGases from different origins.
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