A Low-Temperature-Active Pectate Lyase from a Marine Bacterium for Orange Juice Clarification.

Cold-adapted pectin lyases are particularly useful in the extraction and clarification of freshly squeezed fruit juices at low temperatures, as they effectively reduce juice viscosity and improve light transmittance. With the increasing attention on low-temperature pectinase in industrial applications, the exploration of low-temperature pectinase with novel characteristics has become one of the key focuses of research and development. In this study, a 1026 bp gene, pel1Ba, encoding a 42.7 kDa pectin lyase, was cloned from sediment samples collected from the South China Sea and heterologously expressed in Escherichia coli. The purified Pel1Ba exhibited an optimal temperature of 40 °C and an optimal pH of 10, with a total enzyme activity of 5100 U/mL. Notably, Pel1Ba is a cold-adapted enzyme that retains 80% of its relative activity across the temperature range of 0-40 °C. When 20 U/mL purified Pel1Ba was added to orange juice, the juice volume increased by 43.00% and its clarity improved by 37.80%. Meanwhile, site-directed mutagenesis analysis revealed that the residual enzyme activities of the mutants A230I, F253I, and L292I were increased by 22.5%, 34.4%, and 25.1%, respectively, compared to the wild type. This study concludes that the cold-active pectate lyase Pel1Ba exhibits potential for applications in the food industry.