title: Enhanced sialylation of recombinant erythropoietin in genetically engineered Chinese-hamster ovary cells.
abstract: Sialic acid, the terminal sugar in N-linked complex glycans, is usually found in glycoproteins and plays a major role in determining the circulatory lifespan of glycoproteins. In the present study we attempted to enhance the sialylation of recombinant EPO (erythropoietin) in CHO (Chinese-hamster ovary) cells. To enhance EPO sialylation, we introduced human alpha2,3-ST (alpha2,3-sialyltransferase) and CMP-SAS (CMP-sialic acid synthase) into recombinant human EPO-producing CHO cells. The sialylation of EPO was increased by the expression of alpha2,3-ST alone. Although the co-expression of alpha2,3-ST and CMP-SAS did not further increase sialylation, an increase in the intracellular pool of CMP-sialic acid was noted. On the basis of these observations, it was postulated that the transport capacity of CMP-sialic acid into the Golgi lumen was limited, thereby causing the reduced availability of CMP-sialic acid substrate for sialylation. Therefore, we co-expressed human alpha2,3-ST and CMP-SAS, as well as overexpress Chinese hamster CMP-sialic acid transporter (CMP-SAT) in CHO cells, which produced recombinant human EPO. When alpha2,3-ST, CMP-SAS, and CMP-SAT were overexpressed in CHO cells, there was a corresponding increase in sialylation compared with the co-expression of alpha2,3-ST and CMP-SAS. The present study provides a useful strategy for enhancing the sialylation of therapeutic glycoproteins produced in CHO cells.
authors.affiliation: Department of Biological Sciences, KAIST (Korea Advanced Institute of Science and Technology), 335 Gwahangno, Yuseong-gu, Daejeon 305-701, Republic of Korea.
authors.forename: Yeon Tae
authors.initials: YT
authors.lastname: Jeong
authors.forename: One
authors.initials: O
authors.lastname: Choi
authors.forename: Young Dok
authors.initials: YD
authors.lastname: Son
authors.forename: Seung Yeol
authors.initials: SY
authors.lastname: Park
authors.forename: Jung Hoe
authors.initials: JH
authors.lastname: Kim
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004921:Erythropoietin / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D024861:Genetic Enhancement / Q000379:methods*; D006801:Humans; D019158:N-Acetylneuraminic Acid / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins
chemical_list: D011994:Recombinant Proteins; D004921:Erythropoietin; D019158:N-Acetylneuraminic Acid