1. Novel strategy for expression and characterization of rabies virus glycoprotein.
- Author
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Zhao R, Shan Y, Li M, Lou Z, Feng Y, Huang L, Ren W, Wang P, Sun Y, Sun Y, Su J, Sun H, Hong D, Li Y, Chen R, and Sun L
- Subjects
- Animals, Antigens, Viral genetics, Antigens, Viral immunology, Cloning, Molecular, Cross Protection, Escherichia coli genetics, Escherichia coli metabolism, Gene Expression, Genetic Vectors chemistry, Genetic Vectors metabolism, Humans, Immune Sera chemistry, Immunoglobulin G genetics, Immunoglobulin G metabolism, Immunoglobulin Heavy Chains metabolism, Mice, Protein Engineering methods, Protein Sorting Signals genetics, Rabies virology, Rabies Vaccines administration & dosage, Rabies Vaccines biosynthesis, Rabies Vaccines genetics, Rabies virus genetics, Recombinant Fusion Proteins administration & dosage, Recombinant Fusion Proteins biosynthesis, Viral Envelope Proteins genetics, Viral Envelope Proteins immunology, Antibodies, Neutralizing biosynthesis, Antibodies, Viral biosynthesis, Antigens, Viral administration & dosage, Immunoglobulin Heavy Chains genetics, Rabies prevention & control, Rabies virus immunology, Recombinant Fusion Proteins genetics, Viral Envelope Proteins administration & dosage
- Abstract
Rabies is a fatal zoonosis which could affect all mammals. Glycoprotein (G protein) from the rabies virus plays an important role in the binding of virus to target cells. However, expression of the G protein with native conformation has been a great challenge for many years. In this study, we solved this problem by replacing the original signal peptide of rabies virus G protein with the one from the heavy chain of human IgG. The expression levels of recombinant G protein dramatically increased from a few μg/L to 50 mg/L in the culture supernatants. The identity of the recombinant G protein was confirmed by western blotting using both 6XHis mAb 6E2 and rabies G protein mAb 7G3. The correct conformation of the recombinant G protein was shown by using rabies virus neutralizing antibodies. In addition, the recombinant G protein had immune-reactivities with mice sera raised against rabies vaccines and vice versa. Taken together, our data suggested that by replacing the signal peptide, the expression level of the G protein with native conformation could be significantly improved. This would help the development of a rabies subunit vaccine, structural studies of rabies G protein, elucidation of the signal pathway of RABV infection., (Copyright © 2020 Elsevier Inc. All rights reserved.)
- Published
- 2020
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