公开(公告)号：US08343736B2

公开(公告)日：2013-01-01

申请号：US13391073

申请日：2011-01-18

申请人： Jung Hoe Kim ,  Byoung Hoon Yoon ,  Woo Young Jeon ,  Woo Yong Shim

发明人： Jung Hoe Kim ,  Byoung Hoon Yoon ,  Woo Young Jeon ,  Woo Yong Shim

IPC分类号： C12P21/06 ,  C12N15/19 ,  C12N1/00 ,  C12N15/00

CPC分类号： C12P7/18

摘要： The present invention relates to an efficient production method of xylitol by using the xylitol producing microorganism introduced with arabinose metabolic pathway to inhibit the production of arabitol, the byproduct, and instead to use arabinose only for cell metabolism in xylose/arabinose mixed medium. More precisely, to express efficiently L-arabinose isomerase (araA), L-ribulokinase (araB) and L-ribulose-5-phosphate 4-epimerase (araD) in Candida tropicalis, codon optimization was performed. Then, each gene was inserted in the gene expression cassette containing the glyceraldehyde-3-phosphate dehydrogenase promoter and the selection marker URA3, which was introduced into Candida sp. microorganism. As a result, arabitol, the byproduct interrupting the purification and crystallization of xylitol could be inhibited, making the production method of xylitol of the present invention more efficient. The xylitol producing microorganism introduced with arabinose metabolic pathway of the present invention can be effectively used for the production of xylitol with high productivity by inhibiting the generation of arabitol.

摘要翻译： 本发明涉及通过使用引入了阿拉伯糖代谢途径的木糖醇生产微生物来抑制阿糖醇（副产物）的产生的代谢产物，而是仅将木糖/阿拉伯糖混合培养基中的细胞代谢使用阿拉伯糖，从而有效地生产木糖醇。 更准确地说，为了在热带假丝酵母中有效地表达L-阿拉伯糖异构酶（araA），L-核糖激酶（araB）和L-核酮糖-5-磷酸4-差向异构酶（araD），进行密码子优化。 然后将每个基因插入含有甘油醛-3-磷酸脱氢酶启动子和选择标记URA3的基因表达盒中，将其引入假丝酵母属 微生物。 结果，可以抑制中和木糖醇的纯化和结晶的副产物，从而使本发明的木糖醇的生产方法更有效。 引入本发明的阿拉伯糖代谢途径的木糖醇生产微生物可以通过抑制阿拉伯糖醇的产生而以高生产率有效地用于生产木糖醇。

公开(公告)号：US20090004704A1

公开(公告)日：2009-01-01

申请号：US11771081

申请日：2007-06-29

申请人： Jung Hoe Kim ,  Byoung Sam Ko

发明人： Jung Hoe Kim ,  Byoung Sam Ko

IPC分类号： C12P19/02 ,  C07H1/00 ,  C12N1/20

CPC分类号： C12N1/16 ,  C12P7/18

摘要： Disclosed herein are Xylitol dehydrogenase-inactivated and arabinose reductase-inhibited mutant of Candida tropicalis, a method of producing a high yield of xylitol using the same, and xylitol produced by the method. More specifically, disclosed are a method for producing a high yield of xylitol, in which a high concentration of xylose contained in a biomass hydrolyzate is converted to xylitol using xylitol dehydrogenase-inactivated mutant of Candida tropicalis, without controlling dissolved oxygen to a low level, as well as xylitol produced according to the method. Also disclosed are a xylitol production method, in which the production of byproduct arabitol, which is produced when using a biomass as a substrate and adversely affects the yield of xylitol, is significantly reduced through the use of Candida tropicalis mutant ara-89 (KCTC 11136bp) having an inhibited activity of arabinose reductase converting arabinose to arabitol, thus increasing xylitol productivity, as well as xylitol produced by the method.

摘要翻译： 本文公开了热带假丝酵母的木糖醇脱氢酶失活和阿拉伯糖还原酶抑制突变体，使用其产生高产率的木糖醇的方法，以及通过该方法生产的木糖醇。 更具体地，公开了一种生产高产率的木糖醇的方法，其中将生物质水解产物中所含的高浓度的木糖转化为木糖醇，使用热带假丝酵母的木糖醇脱氢酶灭活的突变体，而不将溶解氧控制在低水平， 以及根据该方法生产的木糖醇。 还公开了一种木糖醇生产方法，其中通过使用热带假丝酵母突变体ara-89（KCTC 11136bp）显着降低在使用生物质作为底物时产生的副产物阿拉伯糖醇的产生并且不利地影响木糖醇的产率 ）具有抑制阿拉伯糖还原酶的活性，其将阿拉伯糖转化成阿糖醇，从而增加木糖醇生产率，以及通过该方法产生的木糖醇。

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。

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。

公开(公告)号：US20120171739A1

公开(公告)日：2012-07-05

申请号：US13391073

申请日：2011-01-18

申请人： Jung Hoe Kim ,  Byoung Hoon Yoon ,  Woo Young Jeon ,  Woo Yong Shim

发明人： Jung Hoe Kim ,  Byoung Hoon Yoon ,  Woo Young Jeon ,  Woo Yong Shim

IPC分类号： C12P7/18 ,  C12N1/19

CPC分类号： C12P7/18

摘要： The present invention relates to an efficient production method of xylitol by using the xylitol producing microorganism introduced with arabinose metabolic pathway to inhibit the production of arabitol, the byproduct, and instead to use arabinose only for cell metabolism in xylose/arabinose mixed medium. More precisely, to express efficiently L-arabinose isomerase (araA), L-ribulokinase (araB) and L-ribulose-5-phosphate 4-epimerase (araD) in Candida tropicalis, codon optimization was performed. Then, each gene was inserted in the gene expression cassette containing the glyceraldehyde-3-phosphate dehydrogenase promoter and the selection marker URA3, which was introduced into Candida sp. microorganism. As a result, arabitol, the byproduct interrupting the purification and crystallization of xylitol could be inhibited, making the production method of xylitol of the present invention more efficient. The xylitol producing microorganism introduced with arabinose metabolic pathway of the present invention can be effectively used for the production of xylitol with high productivity by inhibiting the generation of arabitol.

摘要翻译： 本发明涉及通过使用引入了阿拉伯糖代谢途径的木糖醇生产微生物来抑制阿糖醇（副产物）的产生的代谢产物，而是仅将木糖/阿拉伯糖混合培养基中的细胞代谢使用阿拉伯糖，从而有效地生产木糖醇。 更准确地说，为了在热带假丝酵母中有效地表达L-阿拉伯糖异构酶（araA），L-核糖激酶（araB）和L-核酮糖-5-磷酸4-差向异构酶（araD），进行密码子优化。 然后将每个基因插入含有甘油醛-3-磷酸脱氢酶启动子和选择标记URA3的基因表达盒中，将其引入假丝酵母属 微生物。 结果，可以抑制中和木糖醇的纯化和结晶的副产物，从而使本发明的木糖醇的生产方法更有效。 引入本发明的阿拉伯糖代谢途径的木糖醇生产微生物可以通过抑制阿拉伯糖醇的产生而以高生产率有效地用于生产木糖醇。