In this paper, we report cloning of a pectate lyase gene from Bacillus amyloliquefaciens S6 (pelS6), and biochemical characterization of the recombinant pectate lyase. PelS6 was found to be identical with B. subtilis 168 pel enzyme with 100% amino acid sequence homology. Although these two are genetically very close, they are distinctly different in physiology. pelS6 gene encodes a 421-aa protein with a molecular mass of 65,75 kDa. Enzyme activity increased from 12.8 ± 0.3 to 49.6 ± 0.4 units/mg after cloning. The relative enzyme activity of the recPel S6 ranged from 80% to 100% at pH between 4 and 14. It was quite stable at different temperature values ranging from 15 to 90 °C. The recPEL S6 showed a maximal activity at pH 10 and at 60 °C. 0.5 mM of CaCl2 is the most effective metal ion on the recPEL S6 as demonstrated by its increased relative activity with 473%. recPEL S6 remained stable at - 20 °C for 18 months. In addition recPEL S6 increased juice clarity. This study introduces a novel bacterial pectate lyase enzyme with its characteristic capability of being highly thermostable, thermotolerant, and active over a wide range of pH, meaning that it can work at both acidic and alkaline environments, which are the most preferred properties in the industry.
Keywords: Bacillus amyloliquefaciens; Pectate lyases; pH-thermostable; pH-thermotolerance.