Development of an Innovative Process for High-Temperature Fruit Juice Extraction Using a Novel Thermophilic Endo-Polygalacturonase From Penicillium oxalicum.

Zhong Cheng,Yunlai Chen,Liqin Du,Zhikai Zhang,Liang Xian,Dewu Bi,Qingyan Wang,Dong Chen,Ribo Huang,Yutuo Wei,Jian Lu,Bo Lu

doi:10.3389/fmicb.2020.01200

Zhong Cheng, Yunlai Chen + Show 10 more

Open Access

https://doi.org/10.3389/fmicb.2020.01200

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Abstract

Efficient and cost-effective production of thermophilic endo-polygalacturonase is desirable for industrial fruit juice production, because its application could shorten the processing time and lower the production cost, by eliminating the separate step of pectin degradation. However, no endo-polygalacturonase that both functions well at sufficiently high temperature and can be manufactured economically, has been reported previously. In this study, the cDNA encoding a thermophilic endo-polygalacturonase from Penicillium oxalicum CZ1028, was cloned and over-expressed in Pichia pastoris GS115 and Escherichia coli BL21(DE3). The recombinant proteins PoxaEnPG28B-Pp (from P. pastoris) and PoxaEnPG28B-Ec (from E. coli) were isolated and purified. PoxaEnPG28B-Pp was sufficiently thermostable for potential industrial use, but PoxaEnPG28B-Ec was not. The optimal pH and temperature of PoxaEnPG28B-Pp were pH 5.0 and 65°C, respectively. The enzyme had a low Km of 1.82 g/L and a high Vmax of 77882.2 U/mg, with polygalacturonic acid (PGA) as substrate. The performance of PoxaEnPG28B-Pp in depectinization of papaya, plantain and banana juices at 65°C for 15 min was superior to that of a reported mesophilic endo-polygalacturonase. PoxaEnPG28B-Pp is the first endo-polygalacturonase reported to show excellent performance at high temperature. An innovative process, including a step of simultaneous heat-treatment and depectinization of fruit pulps with PoxaEnPG28B-Pp, is reported for the first time.

Highlights

Fruit juices are popular beverages in most parts of the world, with many different flavors, that contain abundant nutrients and bioactive components, and meet the personal taste preferences of many consumers (Ahmad and Chwee, 2008)
We previously reported a thermo-active endo-PGase EPG4, purified from a newly isolated Penicillium oxalicum CZ1028, which had optimum enzymatic activity at 60–70◦C (Cheng et al, 2016)
The amino acid sequences (EWSGPLLQISGK and WWDGEGSNGGK) of the thermo active endo-PGase EPG4 were found to be identical to two inner sequences (Glu97 to Lys108 and Trp127 to Lys137) of a putative protein from P. oxalicum 114-2

Summary

Introduction

Fruit juices are popular beverages in most parts of the world, with many different flavors, that contain abundant nutrients and bioactive components, and meet the personal taste preferences of many consumers (Ahmad and Chwee, 2008). The industrial production of fruit juices includes four steps. In the raw material treatment step, fresh fruits are peeled, cut and pulped. Comes the heat-treatment step, whereby fruit pulps are heated at 65◦C for 15 min to: (i) inactivate the enzymes that would reduce the quality of the juice (e.g., polyphenol oxidase) (Illera et al, 2018), (ii) accelerate the disruption of the cell structure, to improve the release of cell contents (e.g., polyphenols and anthocyanin pigments) (Marszałek et al, 2017) and (iii) reduce the risk of microbial contamination (Ahmad and Chwee, 2008). In the fourth and final step, the pulp is pressed and the resulting crude fruit juice is clarified, filtered, pasteurized, and packaged (Lozano, 2006)

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