Molecular Characterizing a Novel Alkaline Endo-Pectate Lyase from Paenibacillus borealis and Over-Production in Bioreactor Realized by Constructing the Tandem Expression Cassettes in Host Genome

Alkaline pectate lyases have great prospects in industry fields such as degumming of papermaking and textile, etc. In this study, a novel pectinase PelA from a Paenibacillus borealis strain was molecular characterized and enzymatically defined. This enzyme represents an important cluster divergent from the well-characterized Bacillus pectinase, shows molecularly active under alkaline condition, and has the optimal pH=9.5. It could be clustered as endo-(1,4)-pectate lyase, and break the α-1,4 glycosidic bonds of the polygalacturonic acid by trans-elimination mode. The extra addition of metal ion Ca2+ could not improve the enzyme activity. In order to achieved its high-level secretory expression and improve its economic competence in bioapplication, the gene copy number of PelA in host genome was improved by constructing the tandem PelA gene expression cassettes. After cultivation condition optimization, cell growth monitoring, the recombinant strain carrying the multi-copy pelA gene reached the expression level of 7520 U/mL culture in a bioreactor. This study has fulfilled the high-level secretory expression of an alkaline pectinase, facilitated its industrial bioapplication and also could be a reference for future work on the heterologous expression of target genes.