title: Improved expression of secretory and trimeric proteins in mammalian cells via the introduction of a new trimer motif and a mutant of the tPA signal sequence.
abstract: Ideal immunogenicity in antigens is a prerequisite to eliciting a sufficiently strong immune and memory response via either DNA or protein vaccines. To improve immunogenicity, efforts have focused on high-level expression of target proteins and on maintaining their natural conformations. In the present work, two trimer motifs (MTQ and MTI) were designed and introduced into a plasmid vector with the tissue plasminogen activator signal peptide (tPA-SP). Next, we examined the efficacy and the efficiency of the two motifs as well as the introduction of tPA-SP and its mutant forms, 22P/A and 22P/G, in facilitating the secretory expression of trimeric proteins in mammalian cells. We found that both trimer motifs could produce the target protein in a trimeric form at a high level. Introduction of tPA-SP 22P/A markedly increased the secretory expression level. The combination of the trimer motif, MTQ, and the signal peptide, 22P/A, may serve as a universal mammalian vector for producing trimeric proteins in vaccine development.
authors.affiliation: Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081, China.
authors.forename: Jia-Ye
authors.initials: JY
authors.lastname: Wang
authors.forename: Wen-Ting
authors.initials: WT
authors.lastname: Song
authors.forename: Yan
authors.initials: Y
authors.lastname: Li
authors.forename: Wen-Jiang
authors.initials: WJ
authors.lastname: Chen
authors.forename: Dan
authors.initials: D
authors.lastname: Yang
authors.forename: Guo-Cai
authors.initials: GC
authors.lastname: Zhong
authors.forename: Hai-Zhou
authors.initials: HZ
authors.lastname: Zhou
authors.forename: Cai-Yun
authors.initials: CY
authors.lastname: Ren
authors.forename: Hao-Tong
authors.initials: HT
authors.lastname: Yu
authors.forename: Hong
authors.initials: H
authors.lastname: Ling
mesh_terms: D020816:Amino Acid Motifs / Q000276:immunology*; D000595:Amino Acid Sequence; D004797:Enzyme-Linked Immunosorbent Assay; D057809:HEK293 Cells; D006678:HIV / Q000235:genetics / Q000378:metabolism; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D006801:Humans; D053122:Influenza A Virus, H3N2 Subtype / Q000235:genetics / Q000378:metabolism; D046912:Multiprotein Complexes; D009154:Mutation; D021382:Protein Sorting Signals / Q000235:genetics*; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D010959:Tissue Plasminogen Activator / Q000235:genetics* / Q000276:immunology; D019444:Vaccines, DNA / Q000276:immunology; D022223:Vaccines, Subunit / Q000276:immunology; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000276:immunology / Q000378:metabolism
chemical_list: D019267:Hemagglutinin Glycoproteins, Influenza Virus; D046912:Multiprotein Complexes; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; D022223:Vaccines, Subunit; D054301:gag Gene Products, Human Immunodeficiency Virus; D010959:Tissue Plasminogen Activator