One-step, inexpensive high yield strategy for Candida antarctica lipase A isolation using hydroxyapatite

• Published in: Bioresource Technology
• Main Authors: Dimitrijević, Aleksandra, Veličković, Dušan, Bihelović, Filip, Bezbradica, Dejan, Jankov, Ratko, Milosavić, Nenad
• Format: Article in Journal/Newspaper
• Language: unknown
• Published: Elsevier Sci Ltd, Oxford 2012
• Subjects: Candida antarctica; Lipase; Isolation; Purification; Hydroxyapatite; Triton; Antarc*; Antarctica
• Online Access: http://TechnoRep.tmf.bg.ac.rs/handle/123456789/2201; https://doi.org/10.1016/j.biortech.2011.11.077

Lipase A from Candida antarctica (CAL A) was purified to apparent homogeneity in a single step using hydroxyapatite (HAP) chromatography. CAL A bound to HAP was eluted with 10 mM Na-phosphate buffer, pH 7.0 containing 0.5% Triton X-100. The protocol resulted in a 3.74-fold purification with 94.7% final recovery and 400.83 U/mg specific activity. Silver staining after SDS-PAGE revealed the presence a single band of 45 kDa. The enzyme exhibited a temperature optimum of 60 degrees C, was unaffected by monovalent metal ions, but was destabilized by divalent metal ions (Zn2+, Ca2+, Mg2+, Cu2+, Mn2+) and stimulated by 50 mM Fe2+. Detergents at 0.1% concentrations did not affect lipase activity. Except for Triton X-100, detergent concentrations of 1% had a destabilizing effect.