High yield expression of Lipase A from Candida antarctica in the methylotrophic yeast Pichia pastoris and its purification and characterisation

• Published in: Applied microbiology and biotechnology Vol. 72; no. 5; pp. 931 - 938
• Main Authors: Pfeffer, Jan, Richter, Sven, Nieveler, Jens, Hansen, Carl-Erik, Rhlid, Rachid Bel, Schmid, Rolf D, Rusnak, Monika
• Format: Journal Article
• Language: English
• Published: Berlin Berlin/Heidelberg : Springer-Verlag 01.10.2006; Springer; Springer Nature B.V
• Subjects: Biological and medical sciences; Biotechnology; Candida - enzymology; Candida antarctica; Enzyme Stability; Enzymes; Esters; Fermentation; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Fungal; Hydrogen-Ion Concentration; Kinetics; Lipase - metabolism; Microbiology; Pichia - genetics; Pichia - metabolism; Pichia pastoris; Temperature; Yeasts; Yeast; Purification; candida antarctica; Enzyme; Triacylglycerol lipase; Esterases; Pichia pastoris; Methylotrophy; Carboxylic ester hydrolases; Fungi; Hydrolases; Ascomycetes; Yield; Fungi Imperfecti; Thallophyta
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-006-0400-z

The current investigation focuses on shedding further light on the characteristics of lipase A from Candida antarctica (CalA), which has attracted growing attention in its suitability for industrial applications. CalA was functionally expressed in the methylotrophic yeast Pichia pastoris, purified and characterised. A classical fed-batch process and a semi-continuous process were developed and tested with regard to their yield capacity. Lipase concentrations of 0.88 and 0.55 g l-¹ were obtained using the fed-batch and semi-continuous processes, respectively. The semi-continuous process reaches a total activity of 10,233,000 U and so surpasses the fed-batch process reaching 7,530,000 U. The purified enzyme showed highest activity between 50 and 70 °C at pH 7.0 and a preference for short-chain triglycerides (C4-C8). Significantly reduced activity was observed in the presence of hydrophilic esters.