title: Albumin 3'untranslated region facilitates increased recombinant protein production from Chinese hamster ovary cells.
abstract: Chinese hamster ovary (CHO) cells are used for recombinant protein production in the pharmaceutical industry but there is a need to improve expression levels. In the present work experiments were carried out to test the effectiveness of different 3'untranslated regions (3'UTRs) in promoting production of a naturally secreted luciferase. Seamless cloning was used to produce expression vectors in which Gaussia princeps luciferase coding sequences were linked to the human albumin, immunoglobulin or chymotrypsinogen 3'UTR. Stably transfected CHO cells expressing these constructs were selected. Luciferase activity in the culture medium was increased 2-3-fold by replacing the endogenous 3'UTR with the albumin 3'UTR and decreased by replacement with immunoglobulin or chymotrypsinogen 3'UTR. Replacement of the native 3'UTR with the albumin 3'UTR led to a 10-fold increase in luciferase mRNA levels. Deletion analysis of the albumin 3'UTR showed that loss of nucleotides 1-50, which removed an AU-rich complex stem loop region, caused significant reductions in both luciferase protein expression and luciferase mRNA levels. The results suggest that recombinant protein expression and yield could be improved by the careful selection of appropriate 3'UTR sequences.
authors.affiliation: Institute for Cell and Molecular Biosciences, The Medical School, Newcastle University, Framlington Place, Newcastle upon Tyne, UK.
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Pearson
authors.forename: Siavash
authors.initials: S
authors.lastname: Khazaipoul
authors.forename: Asta
authors.initials: A
authors.lastname: Optun
authors.forename: Ian F
authors.initials: IF
authors.lastname: Pryme
authors.forename: Beate
authors.initials: B
authors.lastname: Stern
authors.forename: John E
authors.initials: JE
authors.lastname: Hesketh
mesh_terms: D020413:3' Untranslated Regions; D000418:Albumins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D001709:Biotechnology / Q000379:methods*; D016466:CHO Cells; D003001:Cloning, Molecular / Q000379:methods; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D020413:3' Untranslated Regions; D000418:Albumins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D008156:Luciferases