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Purification and biochemical characterization of Yarrowia lipolytica LIP2, a lipase of medical interest for the treatment of pancreatic exocrine insufficiency
Author(s) -
Aloulou Ahmed,
Rodriguez Jorge A.,
Puccinelli Delphine,
Mouz Nicolas,
Leblond Yves,
Carriere Frederic
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a609-a
Subject(s) - tributyrin , chemistry , biochemistry , lipase , yeast , yarrowia , enzyme , substrate (aquarium) , triglyceride , fatty acid , chromatography , biology , cholesterol , ecology
The LIP2 lipase from the yeast Y. lipolytica (YLLIP2) was obtained from two genetically modified strains with multi‐copies of the lip2 gene and further purified using gel filtration and cation exchange chromatography. Four YLLIP2 isoforms were identified and subjected to N‐terminal amino‐acid sequencing and mass spectrometry analysis. These isoforms differed in their glycosylation patterns and their molecular masses ranged from 36,874 to 38,481 Da, whereas the polypeptide mass was 33,385 Da. YLLIP2 substrate specificity was investigated using short, medium and long chain triglyceride (TG) at various pH and bile salt (BS) concentrations, and compared with those of human digestive lipases. YLLIP2 was not inhibited by bile salts with any of the substrate tested, and maximum specific activities were found to be 10,760 IU/mg on tributyrin, 16,920 IU/mg on trioctanoin and 12,260 IU/mg on olive oil at pH 6.0. YLLIP2 possesses the same pattern of regioselectivity as human pancreatic lipase, generating 2‐MG and free fatty acids, the lipolysis products absorbed at the intestinal level. YLLIP2 was found to be fairly stable and still active on long chain TG (1590 IU/mg) at pH 4.0, in the presence of BS. YLLIP2 is therefore tailored to act optimally under the physiological conditions pertaining in the gastro‐intestinal tract, and might be an ideal candidate for enzyme replacement therapy for pancreatic exocrine insufficiency.