Combined replacement effects of human modified β-hexosaminidase B and GM2 activator protein on GM2 gangliosidoses fibroblasts
| Autor: | Hitoshi Sakuraba, Youichi Tajima, Chikako Tasaki, Keisuke Kitakaze, Daisuke Tsuji, Takatsugu Hirokawa, Kohji Itoh |
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| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
endocrine system Hex β-hexosaminidase GM2A GM2 activator protein Biophysics Lysosomal storage disease Biology Biochemistry law.invention 03 medical and health sciences 0302 clinical medicine CM conditioned medium Gm2 activator protein law GM2 GM2 ganglioside medicine modB modified HexB LSD lysosomal storage disease GM2A CI-M6PR cation-independent M6P receptor Ganglioside β-hexosaminidase GM2 gangliosidoses Chinese hamster ovary cell SD Sandhoff disease HEXA medicine.disease Molecular biology HEXB carbohydrates (lipids) ERT enzyme replacement therapy 030104 developmental biology TSD Tay-Sachs disease Enzyme replacement therapy Recombinant DNA biology.protein LAMP-1 lysosomal associated membrane protein 1 lipids (amino acids peptides and proteins) M6P mannose-6-phosphate Gm2 gangliosidosis 030217 neurology & neurosurgery Research Article |
| Zdroj: | Biochemistry and Biophysics Reports |
| ISSN: | 2405-5808 |
| DOI: | 10.1016/j.bbrep.2016.04.012 |
| Popis: | GM2 gangliosidoses are autosomal recessive lysosomal storage diseases (LSDs) caused by mutations in the HEXA, HEXB and GM2A genes, which encode the human lysosomal β-hexosaminidase (Hex) α- and β-subunits, and GM2 activator protein (GM2A), respectively. These diseases are associated with excessive accumulation of GM2 ganglioside (GM2) in the brains of patients with neurological symptoms. Here we established a CHO cell line overexpressing human GM2A, and purified GM2A from the conditioned medium, which was taken up by fibroblasts derived from a patient with GM2A deficiency, and had the therapeutic effects of reducing the GM2 accumulated in fibroblasts when added to the culture medium. We also demonstrated for the first time that recombinant GM2A could enhance the replacement effect of human modified HexB (modB) with GM2-degrading activity, which is composed of homodimeric altered β-subunits containing a partial amino acid sequence of the α-subunit, including the GSEP loop necessary for binding to GM2A, on reduction of the GM2 accumulated in fibroblasts derived from a patient with Tay-Sachs disease, a HexA (αβ heterodimer) deficiency, caused by HEXA mutations. We predicted the same manner of binding of GM2A to the GSEP loop located in the modified HexB β-subunit to that in the native HexA α-subunit on the basis of the x-ray crystal structures. These findings suggest the effectiveness of combinational replacement therapy involving the human modified HexB and GM2A for GM2 gangliosidoses. Highlights • Purification of recombinant human GM2A proteins by CHO cell line overexpressing GM2A. • Reduction of GM2 accumulated in GM2A deficiency fibroblasts by GM2A replacement. • Combined effects of modified HexB and GM2A for HexA deficiency fibroblasts. • In silico prediction of molecular interaction between modified HexB and GM2A. |
| Databáze: | OpenAIRE |
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