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Reduction of alpha-Gal expression by relocalizing alpha-galactosidase to the trans-Golgi network and cell surface.
Glycobiology ( IF 4.3 ) Pub Date : 2002-12-04 , DOI: 10.1093/glycob/cwf076
Simon G Taylor 1 , Narin Osman , Ian F C McKenzie , Mauro S Sandrin
Affiliation  

Historically, the most effective means of modifying cell surface carbohydrates has required the intracellular overexpression of glycosyltransferases or glycosidases and is dependent on the enzymes occupying a cellular localization close to the carbohydrate structures they modify. We report on relocalizing the lysosomal resident glycosidase human alpha-galactosidase to other regions of the cell, Golgi and cell surface, where it is in closer proximity for cleaving the carbohydrate structure Galalpha(1,3)Gal. Relocalization of alpha-galactosidase was achieved by using the transmembrane and cytoplasmic domains from the human protein furin, which is known to localize in the trans-Golgi network (TGN) and cell surface. Two chimeric forms of alpha-galactosidase were generated, one directing it to the TGN of the cell and the other to the cell surface, as shown by confocal microscopy. The relocalized enzymes have the ability to cleave terminal alpha-galactose as detected by expression on the cell surface. Furthermore, when expressed as a transgene in mice, the TGN form of alpha-galactosidase was more effective at decreasing cell surface terminal alpha-galactose than was the native lysosomal form. When expressed in conjunction with the alpha1,2fucosyltransferase that also decreases Galalpha(1,3)Gal, the reduction was additive. The ability to relocalize enzymes that modify cell surface carbohydrate structures has far-reaching implications in biology and may be useful in such fields as xenotransplantation and treatment of glycosidase disorders.

中文翻译:

通过将α-半乳糖苷酶重新定位到反式高尔基体和细胞表面来减少α-Gal表达。

历史上,修饰细胞表面碳水化合物的最有效方法要求糖基转移酶或糖苷酶的细胞内过表达,并且依赖于占据它们所修饰的碳水化合物结构附近的细胞定位的酶。我们报告了溶酶体常驻糖苷酶人类α-半乳糖苷酶重新定位到细胞,高尔基体和细胞表面的其他区域,在那里它更接近于裂解碳水化合物结构Galalpha(1,3)Gal。通过使用人类蛋白弗林蛋白酶的跨膜和胞质结构域实现了α-半乳糖苷酶的重新定位,众所周知,弗林蛋白酶位于跨高尔基体(TGN)和细胞表面。产生了两种嵌合形式的α-半乳糖苷酶,一种将其引导至细胞的TGN,另一种将其引导至细胞表面,如共聚焦显微镜所示。重新定位的酶具有切割末端α-半乳糖的能力,如通过在细胞表面上的表达所检测的。此外,当在小鼠中以转基因表达时,α-半乳糖苷酶的TGN形式比天然的溶酶体形式更有效地减少了细胞表面末端的α-半乳糖。当与也降低Galalpha(1,3)Gal的alpha1,2fucosyltransferase一起表达时,这种降低是累加的。重新定位修饰细胞表面碳水化合物结构的酶的能力在生物学中具有深远的意义,并且可能在异种移植和糖苷酶疾病治疗等领域有用。此外,当在小鼠中以转基因表达时,α-半乳糖苷酶的TGN形式比天然的溶酶体形式更有效地减少了细胞表面末端的α-半乳糖。当与也降低Galalpha(1,3)Gal的alpha1,2fucosyltransferase一起表达时,这种降低是累加的。重新定位修饰细胞表面碳水化合物结构的酶的能力在生物学中具有深远的意义,并且可能在异种移植和糖苷酶疾病治疗等领域有用。此外,当在小鼠中以转基因表达时,α-半乳糖苷酶的TGN形式比天然的溶酶体形式更有效地减少了细胞表面末端的α-半乳糖。当与也降低Galalpha(1,3)Gal的alpha1,2fucosyltransferase一起表达时,这种降低是累加的。重新定位修饰细胞表面碳水化合物结构的酶的能力在生物学中具有深远的意义,并且可能在异种移植和糖苷酶疾病治疗等领域有用。减少是累加的。重新定位修饰细胞表面碳水化合物结构的酶的能力在生物学中具有深远的意义,并且可能在异种移植和糖苷酶疾病治疗等领域有用。减少是累加的。重新定位修饰细胞表面碳水化合物结构的酶的能力在生物学中具有深远的意义,并且可能在异种移植和糖苷酶疾病治疗等领域有用。
更新日期:2019-11-01
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