Purification of a Penicillium citrinum lipase by chromatographic processes |
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Authors: | N. Krieger M. A. Taipa E. H. M. Melo J. L. Lima-Filho M. R. Aires-Barros J. M. S. Cabral |
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Affiliation: | Departamento de Química, Universidade Federal do Paraná, Curitiba, Paraná, Brasil, Laboratório de Engenharia Bioquímica, Centro de Engenharia Biológica e Química, Instituto Superior Técnico, 1000 Lisboa, Portugal, PT Departamento de Bioquímica, Universidade Federal de Pernambuco/Laboratório de Imunopatologia Keizo Azami-LIKA, Recife, Pernambuco, Brasil,
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Abstract: | A lipase from a wild strain of Penicillium citrinum was purified by ammonium sulphate precipitation, gel filtration chromatography on a Superose 6 column and hydrophobic interaction chromatography (HIC) on a Phenyl Superose column. The yield and purification factor were 15.2% and 379 fold, respectively. The gel filtration step was efficiently scaled-up in a Superose 6 preparative grade column and after this step, the lipase was recovered in the form of a high molecular weight aggregate. The partial disaggregation of the complex was achieved by HIC and elution with 1.0% (w/v) CHAPS. The lipase produced by Penicillium citrinum forms a dimmer of 63?000 Da, as determined by SDS-PAGE, and it accumulates in the fermentation broth as high molecular weight aggregates (>2?00?000 Da). The analysis of the dimmer showed two subunits with similar molecular weights (31?000–33?000 Da) and isoelectric points (4.8–5.0). |
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