• BMB Reports
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• 제43권8호
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• pp.541-546
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• 2010

We utilized a mammalian expression system to purify and characterize autotaxin (ATX)/lysophospholipase D, an enzyme present in the blood responsible for biosynthesis of lysophosphatidic acid. The human ATX cDNA encoding amino acids 29-915 was cloned downstream of a secretion signal of CD5. At the carboxyl terminus was a thrombin cleavage site followed by the constant domain (Fc) of IgG to facilitate protein purification. The ATX-Fc fusion protein was expressed in HEK293 cells and isolated from conditioned medium of a stable clone by affinity chromatography with Protein A sepharose followed by cleavage with thrombin. The untagged ATX protein was further purified to essential homogeneity by gel filtration chromatography with a yield of approximately 5 mg/liter medium. The purified ATX protein was enzymatically active and biologically functional, offering a useful tool for further biological and structural studies of this important enzyme.

• Molecular & Cellular Toxicology
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• 제3권4호
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• pp.262-266
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• 2007

ENPP2, a 125 kDa secreted lysophopholipase D which originally identified as a tumor-motogen, Autotaxin, enhances cellular locomotion, cell proliferation, angiogenesis and cell survival by generating the signal molecule lysophosphatic acid or sphingosine-1-phosphate. Previous studies have suggested that expression of Autotaxin is associated with invasive phenotype in advanced breast carcinomas. Thus, to determine whether genetic alterations of ENPP2 gene are involved in the development or progression of breast cancer, we analyzed its somatic mutation in 85 breast carcinomas by single-stranded conformational polymorphism and sequencing. Overall, six ENPP2 mutations were found (7.0%), comprising five missense and one nonsense mutation (s). To our knowledge, this is the first report on ENPP2 mutation in breast carcinoma, and the data indicate that ENPP2 is occasionally mutated in breast carcinomas, and suggest that ENPP2 mutation may contribute to the tumor development in some breast carcinomas.