Comparison of strategies for the production of FMDV empty capsids using the Baculovirus Vector System

Authors
Ruiz, Vanesa; Mignaqui, Ana Clara; Nuñez, M.C.; Reytor, Edel; Escribano, José M.; Wigdorovitz, Andres
Publication Year
2014
Language
English
Format
article
Status
Published version
Description
Recombinant FMDV empty capsids have been produced in insect cells and larvae using the baculovirus expression system, although protein yield and efficiency of capsid assembly have been highly variable. In this work, two strategies were compared for the expression of FMDV A/Arg/01 empty capsids: infection with a dual-promoter baculovirus vector coding for the capsid precursor (P12A) and the protease 3C under the control of the polyhedrin and p10 promoters, respectively (BacP12A-3C), or a single-promoter vector coding the P12A3C cassette (BacP12A3C). Expression levels and assembly into empty capsids were analyzed in insect cells and larvae. We observed that the use of the single-promoter vector allowed higher levels of expression both in insect cells and larvae. Recombinant capsid proteins produced by both vectors were recognized by monoclonal antibodies (mAbs) directed against conformational epitopes of FMDV A/Arg/01 and proved to self-assemble into empty capsids (75S) and pentamers (12S) when analyzed by sucrose gradient centrifugation.
Instituto de Virología
Fil: Ruiz, Vanesa. INTA. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Mignaqui, Ana Clara. INTA. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Nuñez, M.C. Universidad Politécnica de Madrid. Parque Científico y Tecnológico. Alternative Gene Expression S.L. (ALGENEX); España
Fil: Reytor, Edel. Universidad Politécnica de Madrid. Parque Científico y Tecnológico. Alternative Gene Expression S.L. (ALGENEX); España
Fil: Escribano, José M. Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA). Departamento de Biotecnología; España
Fil: Wigdorovitz, Andres. INTA. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Source
Molecular Biotechnology 56 (11) : 963–970 (November 2014)
Subject
Enfermedades de los Animales
Fiebre Aftosa
Virus Fiebre Aftosa
Baculovirus
Células
Animal Diseases
Foot and Mouth Disease
Aphthovirus
Cells
Access level
Restricted access
License
Repository
INTA Digital (INTA)
Institution
Instituto Nacional de Tecnología Agropecuaria
OAI Identifier
oai:localhost:20.500.12123/4310