Gene Cloning, Characterization and Transesterification Reactions of Mgl-C255, a Lipolytic Enzyme from <i>Neobacillus thermocopriae</i> C255 Isolated from Ash from Popocatépetl Volcano
2024
Graciela Espinosa-Luna | Aaron S. Bustos-Baena | Rocio Solis-Palacios | Jonathan Lara-Sanchez | Aurelio Espinosa-Honorato | Rosa María Oliart-Ros
Lipases and carboxylesterases are enzymes of biotechnological interest both for their reactions and their specificity. They have wide-ranging applications in the food, pharmaceuticals, biodiesel synthesis, and bioremediation industries. For that reason, the strain <i>Neobacillus thermocopriae</i> C255 was isolated from ash from Popocatepetl volcano and studied as a new source of lipolytic enzymes. It was identified using 16S ribosomal RNA and flagellar protein FliF sequence homology, yielding 100% identity. From the sequencing of its genome, an enzyme with lipolytic activity, classified as a monoacylglycerol lipase, and named Mgl-C255, was cloned in <i>E. coli</i> BL21, and then expressed, biochemically characterized, and tested via transesterification reactions with alcohols and monosaccharides. Based on its sequence and structure, it was placed within family V, having a catalytic triad of S90-D207-H237. Biochemical characterization showed its highest activity at 40 °C, pH 7.5 to 8.5, with C-2 length substrate preference. No metal ions or inhibitors influenced lipolytic activity, except for PMSF, SDS, Cu<sup>−2</sup>, and Hg<sup>−2</sup>. Mgl-C255 retained about 50% of its activity in non-polar solvents and showed synthetic activity in organic solvents, making it a good candidate for studying its catalytic potential and selectivity.
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