Coexpression of novel furin-resistant LPL variants with lipase maturation factor 1 enhances LPL secretion and activity
Autor: | Ellis M. Pearson, Ming Jing Wu, Aspen R. Gutgsell, Alan T. Remaley, Anna Wolska, Saskia B. Neher, Benjamin S. Roberts |
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Rok vydání: | 2018 |
Předmět: |
Male
0301 basic medicine Glycan Blotting Western QD415-436 Biochemistry law.invention Mice 03 medical and health sciences Endocrinology trafficking In vivo law protein purification Protein purification Methods Animals Humans Secretion Furin chemistry.chemical_classification Lipoprotein lipase biology Chemistry digestive oral and skin physiology Membrane Proteins nutritional and metabolic diseases Biological Transport Cell Biology Lipoprotein Lipase 030104 developmental biology Enzyme biology.protein Recombinant DNA lipids (amino acids peptides and proteins) |
Zdroj: | Journal of Lipid Research, Vol 59, Iss 12, Pp 2456-2465 (2018) |
ISSN: | 0022-2275 |
Popis: | LPL is a secreted enzyme that hydrolyzes triglycerides from circulating lipoproteins. Individuals lacking LPL suffer from severe hypertriglyceridemia, a risk factor for acute pancreatitis. One potential treatment is to administer recombinant LPL as a protein therapeutic. However, use of LPL as a protein therapeutic is limited because it is an unstable enzyme that is difficult to produce in large quantities. Furthermore, these considerations also limit structural and biochemical studies that are needed for large-scale drug discovery efforts. We demonstrate that the yield of purified LPL can be dramatically enhanced by coexpressing its maturation factor, LMF1, and by introducing novel mutations into the LPL sequence to render it resistant to proteolytic cleavage by furin. One of these mutations introduces a motif for addition of an N-linked glycan to the furin-recognition site. Furin-resistant LPL has previously been reported, but is not commonly used. We show that our modifications do not adversely alter LPL's enzymatic activity, stability, or in vivo function. Together, these data show that furin-resistant LPL is a useful reagent for both biochemical and biomedical studies. |
Databáze: | OpenAIRE |
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