EVALUATION OF ACTIVITY AND MASS SPECTROMETRIC CHARACTERIZATION OF TWO PARTIALLY PURIFIED THERMO-TOLE
Fungal infestations of groundnut seeds were screened for thermo-tolerant lipases with stability over a wide pH range and temperature, desirable for industrial applications. DNA sequencing of ITS 1, 5.8S and ITS 2 regions indicated the isolate closer to Aspergillus niger with 99% identity and therefore designated as A. niger GN1 in this study. The pellet obtained from culture extract subjected to 65% ammonium sulphate precipitation was suspended in Tris-buffer and assayed for lipase activity. Two fractions of lipase (1 and 2) could be extracted at pH 4 and pH 9 respectively. Both lipase 1 and 2 fractions were characterized by LC-MS/MS spectroscopic analysis. The relative and residual activities of the enzyme fractions were high in the temperature range of 60 – 80°C and over a pH of 4 & 8 for lipase 1 and 2-6 for lipase 2 fractions. LC-MS/MS confirmed the presence of Lipase 1 (~32 kDa) and Lipase 2 (~30 kDa) fractions with 4 and 2 unique peptides respectively.
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