Heterologous Expression and Characterization of Thermostable Lipases from Geobacillus Thermoleovorans PPD2 Through Escherichia Coli

Anita H. Permana^1,4, Fida Madayanti Warganegara¹, Deana Wahyuningrum², Made Puspasari Widhiastuty¹ and Akhmaloka^1,3

¹Biochemistry Research Group, Faculty of Mathematics and Natural Science, Institut Teknologi Bandung, Indonesia.

²Organic Chemistry Research Group, Faculty of Mathematics and Natural Science, Institut Teknologi Bandung, Indonesia.

³Department of Chemistry, Faculty of Science and Computer, Universitas Pertamina, Indonesia.

⁴Study Program of Quality Assurance of Food Industry, Politeknik AKA Bogor, Indonesia.

Corresponding Author  E-mail: loka@chem.itb.ac.id

DOI : http://dx.doi.org/10.13005/bbra/2545

ABSTRACT: Heterologous expression and purification of thermostable lipase from Geobacillus thermoleovorans PPD2 had been carried out through Escherichia coli BL21 as host. Two bands obtained showed lipolytic activity with the size at around 51 (LipA) and 43 (LipB) kDa, respectively. The activities were identified by zymogram analysis, while the control protein from Escherichia coli BL21(DE3) do not show any lipolytic activity. Purification of crude extract using chromatography affinity Ni-NTA resulted one dominant band of LipA, meanwhile LipB did not appeared on the gel. Another purification for LipB was carried out by acetone fractionation. Both of LipA and LipB showed high activity toward medium chain length substrates, with optimum activity at 50^oC and pH 8.5. The activities of LipA and LipB showed tolerance toward short chain alcohols, such as methanol, ethanol, n-propanol, and isopropanol.

KEYWORDS: Acetone Fractionation Geobacillus Thermoleovorans PPD2; Thermostable Lipase; Recombinant Protein; Ni-NTA Affinity Chromatography;

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