Characterization of an extracellular lipase encoded by LIP2 in Yarrowia lipolytica.
We isolated the LIP2 gene from the lipolytic yeast Yarrowia lipolytica. It was found to encode a 334-amino-acid precursor protein. The secreted lipase is a 301-amino-acid glycosylated polypeptide which is a member of the triacylglycerol hydrolase family (EC 3.1.1.3). The Lip2p precursor protein is processed by the KEX2-like endoprotease encoded by XPR6. Deletion of the XPR6 gene resulted in the secretion of an active but less stable proenzyme. Thus, the pro region does not inhibit lipase secretion and activity. However, it does play an essential role in the production of a stable enzyme. Processing was found to be correct in LIP2(A) (multiple LIP2 copy integrant)-overexpressing strains, which secreted 100 times more activity than the wild type, demonstrating that XPR6 maturation was not limiting. No extracellular lipase activity was detected with the lip2 knockout (KO) strain, strongly suggesting that extracellular lipase activity results from expression of the LIP2 gene. Nevertheless, the lip2 KO strain is still able to grow on triglycerides, suggesting an alternative pathway for triglyceride utilization in Y. lipolytica.
Pignède
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Laboratoire Mayoly Spindler, Service Recherche, 78401 Chatou Cedex, France.
Wang
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Fudalej
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Gaillardin
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Seman
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Nicaud
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JM
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins; D001483:Base Sequence; D003470:Culture Media; D004271:DNA, Fungal; D005656:Fungal Proteins; D015870:Gene Expression; D005800:Genes, Fungal; D005822:Genetic Vectors; D006031:Glycosylation; D008049:Lipase / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D011817:Rabbits; D004718:Saccharomycetales / Q000201:enzymology* / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism; D014280:Triglycerides / Q000378:metabolism
D001426:Bacterial Proteins; D003470:Culture Media; D004271:DNA, Fungal; D005656:Fungal Proteins; D014280:Triglycerides; C408971:LIP2 protein, Yarrowia lipolytica; D008049:Lipase; D012697:Serine Endopeptidases; C087588:XPR6 protein, Yarrowia lipolytica