Isolation and Partial Characterization of a Bacterial Thermostable Polymethyl Galacturonase from a Newly Isolated Bacillus sp. strain BR1390

Abstract

Background: Pectinases are pectin degrading class of enzymes including polygalacturonase (PG), polymethyl galacturonase (PMG), pectate lyase (PEL), and pectin esterase (PE) that are commonly used in processes involving the degradation of plant materials, such as speeding up the extraction of fruit juices. Objectives: A highly methylated pectin degrading bacterium from soil covered with fruit waste was isolated and its extracellular pectinase as a novel polymethyl galacturonase was characterized. Materials and Methods: Pectin-degrading microorganism screening was performed using agar plates containing pectin as the sole carbon source. The biochemical studies were used to characterize the enzyme. Results: Bacterium with greater PMG activity was a Bacillus sp. based on 16S rDNA sequence analysis and named as a Bacillus sp. strain BR1390. Two steps column chromatography showed a dimeric protein with apparent molecular masses of 104 and 56 kDa, evident by gel filtration and SDS-PAGE. Substrate specificity analysis using various polygalacturonic acid compounds revealed its polymethyl galacturonase (PMG, EC 3.2.1.-) activity. Biochemical studies represent the thermophilic characteristics and reasonable pH stability of the polymethyl galacturonase when using pectin as substrate. The PMG activity significantly enhanced in the presence of most divalent cations such as Ca2+ and Mg2+, but Hg2+ and Fe3+ served as strong inhibitor. Conclusions: Overall, regarding to have suitable activity in acidic conditions and high operational stability of the purified pectinase, the introduced PMG can be an ideal functional substitute for applications in the fruit juice industry, especially in citrus fruits extraction and clarification.