Unveiling six novel CALB-like lipases using genome-centric and patent-driven prospection

Area of Science:

• Biotechnology
• Enzymology
• Bioinformatics

Background:

• Lipases are crucial biocatalysts with broad industrial applications.
• High costs and extensive patenting of enzymes like Candida antarctica lipase B (CALB) limit their widespread use.
• Identifying novel, patent-free lipases is essential for advancing biocatalysis.

Purpose of the Study:

• To identify and characterize novel CALB-like lipases free from patent protection.
• To explore variations in catalytic pocket structure and substrate specificity among these new enzymes.
• To provide new biocatalyst options for diverse biotechnological applications.

Main Methods:

• Genomic searches across nearly 7000 fungal genomes to identify CALB homolog candidates.
• Phylogenetic analysis and intellectual property screening to filter for patent-free sequences.
• Cloning, production, and characterization of selected CALB-like lipases, including substrate specificity assays.

Main Results:

• Six patent-free CALB-like lipases were identified and produced.
• Enzymes exhibited variations in catalytic pocket amino acids and substrate preferences.
• One lipase showed preference for C4 p-nitrophenyl ester due to lower pocket entrance hydrophobicity; another displayed no preference with a wider, more polar pocket.

Conclusions:

• New patent-free lipases with conserved catalytic elements but diverse substrate specificities were discovered.
• These enzymes represent valuable starting points for developing innovative biocatalysts.
• The findings support the implementation of novel lipases in various biotechnological sectors.