A Novel Endo-Polygalacturonase from Penicillium oxalicum: Gene Cloning, Heterologous Expression and Its Use in Acidic Fruit Juice Extraction

Abstract

An endo-polygalacturonase (endo-PGase) exhibiting excellent performance during acidic fruit juice production would be highly attractive to the fruit juice industry. However, candidate endo-PGases for this purpose have rarely been reported. In this study, we expressed a gene from Penicillium oxalicum in Pichia pastoris . The recombinant enzyme PoxaEnPG28C had an optimal enzyme activity at pH 4.5 and 45°C and was stable at pH 3.0–6.5 and < 45°C. The enzyme had a specific activity of 4,377.65 ± 55.37 U/mg towards polygalacturonic acid, and the K m and V max values of PoxaEnPG28C were calculated as 1.64 g/l and 6127.45 U/mg, respectively. PoxaEnPG28C increased the light transmittance of orange, lemon, strawberry and hawthorn juice by 13.9 ± 0.3%, 29.4 ± 3.8%, 95.7 ± 10.2% and 79.8 ± 1.7%, respectively; it reduced the viscosity of the same juices by 25.7 ± 1.6%, 52.0 ± 4.5%, 48.2 ± 0.7% and 80.5 ± 2.3%, respectively, and it increased the yield of the juices by 24.5 ± 0.7%, 12.7 ± 2.2%, 48.5 ± 4.2% and 104.5 ± 6.4%, respectively. Thus, PoxaEnPG28C could be considered an excellent candidate enzyme for acidic fruit juice production. Remarkably, fruit juice production using hawthorn as an material was reported for the first time.

Heterologous Expression and Characterization of a Novel Exo-Polygalacturonase from Aspergillus fumigatus Af293 and Its Application in Juice Extraction

“…The gene was amplified by the primers [5´-GCGC GAATTC GCTCCTAACCAACCAATTCA-3´ (EcoRI was underlined) and 5´-ATAT GCGGCCGC TTAGTGTGGAACGAAGGTAC-3´ (NotI was underlined) ], and the amplification procedure was followed according to a previous report [ 18 ]. Then, the gene was digested with EcoRI and NotI, and inserted into the vector pPIC9K.…”