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1. 发明授权

US08940876B2 Method for preparing recombinant glycoproteins with high sialic acid content 有权
标题翻译：制备高唾液酸含量的重组糖蛋白的方法
公开(公告)号：US08940876B2
公开(公告)日：2015-01-27
申请号：US13390671
申请日：2011-02-01
申请人： Jung Hoe Kim , Young Dok Son , Jin Young Hwang , Yeon Tae Jeong
发明人： Jung Hoe Kim , Young Dok Son , Jin Young Hwang , Yeon Tae Jeong
IPC分类号： C07K14/505 , C07K14/705 , C07K14/52 , C12P21/00 , C12N9/90 , C12N9/12 , C12N9/10
CPC分类号： C07K14/505 , C07K14/524 , C07K14/705 , C12N9/1081 , C12N9/1205 , C12N9/90 , C12P21/005 , C12Y501/03014
摘要： The present disclosure relates to a method for preparing recombinant glycoproteins with high sialic acid content. More specifically, for UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE/MNK) enzyme where point mutation was induced by substituting arginine at position 263 by leucine only or by further substituting arginine at position 266 by glutamine, epimerase activity is constantly maintained, and overexpressed cells thereof experience an increase in intracellular cytidine monophosphate (CMP)-sialic acid content, irrespective of CMP-sialic acid concentration. Particularly, since in an glycoprotein (such as, erythropoietin and thrombopoietin)-producing host cell where point mutationinduced GNE/MNK, human alpha-2,3-sialyltransferase and a CMP-sialic acid transporter gene are simultaneously overexpressed, intracellular content of CMP-sialic acid and sialic acid in glycoprotein increases in cells, overexpression in a host cell producing a sialylated recombinant glycoprotein the three genes above may be useful for preparing glycoprotein with increased sialic acid content.
摘要翻译：本公开涉及一种制备高唾液酸含量的重组糖蛋白的方法。更具体地，对于通过仅通过亮氨酸取代位置263处的精氨酸或通过用谷氨酰胺进一步取代位置266处的精氨酸而诱发点突变的UDP-GlcNAc 2-差向异构酶/ ManNAc激酶（GNE / MNK）酶，不断维持差向异构酶活性，并且其过表达的细胞经历细胞内胞苷酸单磷酸（CMP） - 唾液酸含量的增加，而不考虑CMP-唾液酸浓度。特别是，由于在产生点突变诱导的GNE / MNK的糖蛋白（例如促红细胞生成素和血小板生成素）中，人α-2,3-唾液酸转移酶和CMP-唾液酸转运蛋白基因同时过表达，因此CMP- 糖蛋白中的唾液酸和唾液酸在细胞中增加，在产生唾液酸化重组糖蛋白的宿主细胞中过表达，上述三种基因可能用于制备具有增加的唾液酸含量的糖蛋白。

2. 发明申请

US20120149874A1 METHOD FOR PREPARING RECOMBINANT GLYCOPROTEINS WITH HIGH SIALIC ACID CONTENT 有权
标题翻译：用高碱性内容物制备重组蛋白糖的方法
公开(公告)号：US20120149874A1
公开(公告)日：2012-06-14
申请号：US13390671
申请日：2011-02-01
申请人： Jung Hoe Kim , Young Dok Son , Jin Young Hwang , Yeon Tae Jeong
发明人： Jung Hoe Kim , Young Dok Son , Jin Young Hwang , Yeon Tae Jeong
IPC分类号： C07K14/435 , C12N9/18 , C12N9/10 , C07K14/575 , C12N15/81 , C07K14/745 , C12N15/85 , C12N1/19 , C12N5/10 , C12P21/00 , C07K14/755
CPC分类号： C07K14/505 , C07K14/524 , C07K14/705 , C12N9/1081 , C12N9/1205 , C12N9/90 , C12P21/005 , C12Y501/03014
摘要： The present disclosure relates to a method for preparing recombinant glycoproteins with high sialic acid content. More specifically, for UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE/MNK) enzyme where point mutation was induced by substituting arginine at position 263 by leucine only or by further substituting arginine at position 266 by glutamine, epimerase activity is constantly maintained, and overexpressed cells thereof experience an increase in intracellular cytidine monophosphate (CMP)-sialic acid content, irrespective of CMP-sialic acid concentration. Particularly, since in an glycoprotein(such as, erythropoietin and thrombopoietin)-producing host cell where point mutationinduced GNE/MNK, human alpha-2,3-sialyltransferase and a CMP-sialic acid transporter gene are simultaneously overexpressed, intracellular content of CMP-sialic acid and sialic acid in glycoprotein increases in cells, overexpression in a host cell producing a sialylated recombinant glycoprotein the three genes above may be useful for preparing glycoprotein with increased sialic acid content.
摘要翻译：本公开涉及一种制备高唾液酸含量的重组糖蛋白的方法。更具体地，对于通过仅通过亮氨酸取代位置263处的精氨酸或通过用谷氨酰胺进一步取代位置266处的精氨酸而诱发点突变的UDP-GlcNAc 2-差向异构酶/ ManNAc激酶（GNE / MNK）酶，不断维持差向异构酶活性，并且其过表达的细胞经历细胞内胞苷酸单磷酸（CMP） - 唾液酸含量的增加，而不考虑CMP-唾液酸浓度。特别是，由于在产生点突变诱导的GNE / MNK的糖蛋白（例如促红细胞生成素和血小板生成素）中，人α-2,3-唾液酸转移酶和CMP-唾液酸转运蛋白基因同时过表达，因此CMP- 糖蛋白中的唾液酸和唾液酸在细胞中增加，在产生唾液酸化重组糖蛋白的宿主细胞中过表达，上述三种基因可能用于制备具有增加的唾液酸含量的糖蛋白。

3. 发明申请

US20130203617A1 Glycoprotein Analysis Kit and Use Thereof 审中-公开
标题翻译：糖蛋白分析试剂盒及其用途
公开(公告)号：US20130203617A1
公开(公告)日：2013-08-08
申请号：US13695197
申请日：2011-04-29
申请人： Hak Sung Kim , Sun Young Park , Sang Chul Lee , Eun Kyoung Kim , Sung Ho Kim , Wan Jung Kim , Yong Tae Kim , Young Dok Son , Jung Hoe Kim
发明人： Hak Sung Kim , Sun Young Park , Sang Chul Lee , Eun Kyoung Kim , Sung Ho Kim , Wan Jung Kim , Yong Tae Kim , Young Dok Son , Jung Hoe Kim
IPC分类号： G01N33/74 , C12N15/85 , G01N33/68
CPC分类号： G01N33/746 , C12N15/85 , G01N33/6803 , G01N33/6842 , G01N2400/02
摘要： Disclosed are a glycoprotein analysis kit and use thereof. The glycoprotein analysis kit comprises a fluorescently labeled antibody, a fluorescently labeled biomaterial, and a support, and is used in a dual probing method for analyzing the content of a glycoprotein and profiling the oligosaccharide chain, simultaneously, and a method for selecting a single cell producing the glycoprotein having a desired glycosylation pattern. Also, a single cell producing the glycoprotein having a desired glycosylation pattern is provided.
摘要翻译：公开了糖蛋白分析试剂盒及其用途。糖蛋白分析试剂盒包含荧光标记的抗体，荧光标记的生物材料和载体，并用于同时用于分析糖蛋白含量和分析寡糖链的双重探测方法，以及选择单细胞的方法产生具有所需糖基化模式的糖蛋白。此外，提供了产生具有所需糖基化模式的糖蛋白的单细胞。

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