A quantitative assay measuring the function of lipase maturation factor 1
Newly synthesized lipoprotein lipase (LPL) and related members of the lipase gene family require an endoplasmic reticulum maturation factor for attainment of enzyme activity. This factor has been identified as lipase maturation factor 1 (Lmf1), and mutations affecting its function and/or expression...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2009-11-01
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| Series: | Journal of Lipid Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022227520306921 |
| _version_ | 1819293510869712896 |
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| author | Fen Yin Mark H. Doolittle Miklós Péterfy |
| author_facet | Fen Yin Mark H. Doolittle Miklós Péterfy |
| author_sort | Fen Yin |
| collection | DOAJ |
| description | Newly synthesized lipoprotein lipase (LPL) and related members of the lipase gene family require an endoplasmic reticulum maturation factor for attainment of enzyme activity. This factor has been identified as lipase maturation factor 1 (Lmf1), and mutations affecting its function and/or expression result in combined lipase deficiency (cld) and hypertriglyceridemia. To assess the functional impact of Lmf1 sequence variations, both naturally occurring and induced, we report the development of a cell-based assay using LPL activity as a quantitative reporter of Lmf1 function. The assay uses a cell line homozygous for the cld mutation, which renders endogenous Lmf1 nonfunctional. LPL transfected into the mutant cld cell line fails to attain activity; however, cotransfection of LPL with wild-type Lmf1 restores its ability to support normal lipase maturation. In this report, we describe optimized conditions that ensure the detection of a complete range of Lmf1 function (full, partial, or complete loss of function) using LPL activity as the quantitative reporter. To illustrate the dynamic range of the assay, we tested several novel mutations in mouse Lmf1. Our results demonstrate the ability of the assay to detect and analyze Lmf1 mutations having a wide range of effects on Lmf1 function and protein expression. |
| first_indexed | 2024-12-24T04:11:35Z |
| format | Article |
| id | doaj.art-e5164f45b95249c7b5906bd93ecb220b |
| institution | Directory Open Access Journal |
| issn | 0022-2275 |
| language | English |
| last_indexed | 2024-12-24T04:11:35Z |
| publishDate | 2009-11-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Lipid Research |
| spelling | doaj.art-e5164f45b95249c7b5906bd93ecb220b2022-12-21T17:16:03ZengElsevierJournal of Lipid Research0022-22752009-11-01501122652269A quantitative assay measuring the function of lipase maturation factor 1Fen Yin0Mark H. Doolittle1Miklós Péterfy2Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095; Department of Medicine, VA Greater Los Angeles Healthcare System, Los Angeles, CA 90073; Medical Genetics Institute, Cedars-Sinai Medical Center, Los Angeles, CA 90048To whom correspondence should be addressed; Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095; Department of Medicine, VA Greater Los Angeles Healthcare System, Los Angeles, CA 90073To whom correspondence should be addressed; Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095; Department of Medicine, VA Greater Los Angeles Healthcare System, Los Angeles, CA 90073; Medical Genetics Institute, Cedars-Sinai Medical Center, Los Angeles, CA 90048Newly synthesized lipoprotein lipase (LPL) and related members of the lipase gene family require an endoplasmic reticulum maturation factor for attainment of enzyme activity. This factor has been identified as lipase maturation factor 1 (Lmf1), and mutations affecting its function and/or expression result in combined lipase deficiency (cld) and hypertriglyceridemia. To assess the functional impact of Lmf1 sequence variations, both naturally occurring and induced, we report the development of a cell-based assay using LPL activity as a quantitative reporter of Lmf1 function. The assay uses a cell line homozygous for the cld mutation, which renders endogenous Lmf1 nonfunctional. LPL transfected into the mutant cld cell line fails to attain activity; however, cotransfection of LPL with wild-type Lmf1 restores its ability to support normal lipase maturation. In this report, we describe optimized conditions that ensure the detection of a complete range of Lmf1 function (full, partial, or complete loss of function) using LPL activity as the quantitative reporter. To illustrate the dynamic range of the assay, we tested several novel mutations in mouse Lmf1. Our results demonstrate the ability of the assay to detect and analyze Lmf1 mutations having a wide range of effects on Lmf1 function and protein expression.http://www.sciencedirect.com/science/article/pii/S0022227520306921combined lipase deficiencyhypertriglyceridemialipoprotein lipasegenetic complementation |
| spellingShingle | Fen Yin Mark H. Doolittle Miklós Péterfy A quantitative assay measuring the function of lipase maturation factor 1 Journal of Lipid Research combined lipase deficiency hypertriglyceridemia lipoprotein lipase genetic complementation |
| title | A quantitative assay measuring the function of lipase maturation factor 1 |
| title_full | A quantitative assay measuring the function of lipase maturation factor 1 |
| title_fullStr | A quantitative assay measuring the function of lipase maturation factor 1 |
| title_full_unstemmed | A quantitative assay measuring the function of lipase maturation factor 1 |
| title_short | A quantitative assay measuring the function of lipase maturation factor 1 |
| title_sort | quantitative assay measuring the function of lipase maturation factor 1 |
| topic | combined lipase deficiency hypertriglyceridemia lipoprotein lipase genetic complementation |
| url | http://www.sciencedirect.com/science/article/pii/S0022227520306921 |
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