The C-terminal N-glycosylation sites of the human alpha1,3/4-fucosyltransferase III, -V, and -VI (hFucTIII, -V, adn -VI) are necessary for the expression of full enzyme activity.
The C-terminal N-glycosylation sites of the human alpha1,3/4-fucosyltransferase III, -V, and -VI (hFucTIII, -V, adn -VI) are necessary for the expression of full enzyme activity.
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人 α1,3/4-岩藻糖基转移酶 III、-V 和 -VI (hFucTIII、-V、adn -VI) 的 C 端 N-糖基化位点对于表达完整的酶活性是必需的。
作者：
Lise;Uffe Birk Jensen;Peter Bross;T. Ørntoft
α 1，3/4-岩藻糖基转移酶参与岩藻糖基化细胞表面糖缀合物的合成。人α 1，3/4-岩藻糖基转移酶III、-V和-VI（hFucTIII、-V和-VI）含有两个保守的C-末端N-糖基化位点（hFucTIII：Asn 154和Asn 185; hFucTV：Asn 167和Asn 198; hFucTVI：Asn 153和Asn 184）。在本研究中，我们分析了这些潜在的N-糖基化位点的功能作用，主要侧重于hFucTIII中的位点。衣霉素处理完全消除hFucTIII酶活性，而栗精胺处理使hFucTIII酶活性减少至天然酶活性的约40%。为了进一步分析hFucTIII、-V和-VI中保守的N-糖基化位点的作用，我们制备了一系列突变基因组DNA，其中潜在C-末端N-糖基化位点中的天冬酰胺残基被谷氨酰胺取代。随后，hFucTIII、-V和-VI野生型和突变体在COS-7细胞中表达。所有的突变体表现出较低的酶活性比野生型和消除个别网站上的活动有不同的影响。突变不影响细胞中突变体的蛋白质水平，但如预测的那样降低了分子量。hFucTIII的动力学分析显示，Asn 185处糖基化的缺乏没有改变寡糖受体和核苷酸糖供体的Km值。本研究表明，hFucTIII，-V，和-VI需要在两个保守的C-末端N-糖基化位点的N-糖基化的完整的酶活性的表达。
The alpha1,3/4-fucosyltransferases are involved in the synthesis of fucosylated cell surface glycoconjugates. Human alpha1,3/4-fucosyltransferase III, -V, and -VI (hFucTIII, -V, and -VI) contain two conserved C-terminal N-glycosylation sites (hFucTIII: Asn154 and Asn185; hFucTV: Asn167 and Asn198; and hFucTVI: Asn153 and Asn184). In the present study, we have analyzed the functional role of these potential N-glycosylation sites, laying the main emphasis on the sites in hFucTIII. Tunicamycin treatment completely abolished hFucTIII enzyme activity while castanospermine treatment diminished hFucTIII enzyme activity to approximately 40% of the activity of the native enzyme. To further analyze the role of the conserved N-glycosylation sites in hFucTIII, -V, and -VI, we made a series of mutant genomic DNAs in which the asparagine residues in the potential C-terminal N-glycosylation sites were replaced by glutamine. Subsequently, the hFucTIII, -V, and -VI wild type and the mutants were expressed in COS-7 cells. All the mutants exhibited lower enzyme activity than the wild type and elimination of individual sites had different effects on the activity. The mutations did not affect the protein level of the mutants in the cells, but reduced the molecular mass as predicted. Kinetic analysis of hFucTIII revealed that lack of glycosylation at Asn185 did not change the Km values for the oligosaccharide acceptor and the nucleotide sugar donor. The present study demonstrates that hFucTIII, -V, and -VI require N-glycosylation at the two conserved C-terminal N-glycosylation sites for expression of full enzyme activity.
DOI：
10.1016/s0021-9258(18)42684-1
发表时间：
1992-03
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
O. Ben-Zeev;M. Doolittle;Richard C. Davis;John Elovson;M. Schotz
通讯作者：
O. Ben-Zeev;M. Doolittle;Richard C. Davis;John Elovson;M. Schotz
脂蛋白脂肪酶和肝脂肪酶：天冬酰胺连接的糖基化在功能酶表达中的作用。
影响因子：
6.5
作者：
Ben-Zeev,O;Stahnke,G;Liu,G;Davis,RC;Doolittle,MH
通讯作者：
Doolittle,MH
影响因子：
56.9
作者：
MUECKLER, M;CARUSO, C;LODISH, HF
通讯作者：
LODISH, HF