Thermostable lipase from Geobacillus sp. Iso5: bioseparation, characterization and native structural studies

Area of Science:

• Biochemistry
• Enzymology
• Extremophile Research

Background:

• Extremophilic enzymes, particularly lipases, are valuable for industrial applications due to their stability under harsh conditions.
• Geobacillus species are known sources of robust enzymes, but characterization of their lipases is ongoing.

Purpose of the Study:

• To purify and characterize a novel extracellular thermoalkaline lipase from Geobacillus sp. Iso5.
• To determine the enzyme's optimal activity, stability, and structural properties.

Main Methods:

• Enzyme purification using ultrafiltration and chromatography (agarose, Phenyl Sepharose HIC).
• Molecular weight determination via SDS-PAGE and MALDI-TOF MS/MS.
• Activity and stability assays at various temperatures and pH.
• Enzyme inhibition/promotion studies with various ions and chemicals.
• Secondary structure analysis using Circular Dichroism, FTIR, and Raman Spectroscopy.

Main Results:

• Purification yielded an 8.7-fold increase with 6.2% yield.
• The enzyme is a 47 kDa monomer, optimally active at 70°C and pH 8.0.
• High thermostability observed, retaining >90% activity at 70°C for 2 hours.
• Activity was enhanced by Ca(2+) and Mg(2+), and inhibited by HgCl2, PMSF, DTT, K(+), Co(2+), and Zn(2+).
• Secondary structure analysis revealed 36% α-helix and 64% β-sheet content.

Conclusions:

• Geobacillus sp. Iso5 produces a highly stable thermoalkaline lipase with potential industrial applications.
• The enzyme's characteristics, including its secondary structure, provide insights into its thermostability.
• Further research could explore its specific applications in detergents, food processing, or bioremediation.