Expression of equine morbillivirus (EMV) matrix and fusion proteins and their evaluation as diagnostic reagents

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Abstract
Summary. Full-length cDNA clones coding for the matrix (M) and fusion (F) proteins of equine morbillivirus (EMV) were isolated by RT-PCR, and expressed in Escherichia coli using two different expression systems. Western blot analysis indicated that the M and F proteins, expressed either by itself or as fusion proteins with glutathione S-transferase (GST), were insoluble and degraded after expression. Analysis of the degradation pattern of recombinant M protein suggested that the N-terminus of the matrix protein might be more stable and antigenic than the C-terminal region. Therefore a third system was used to express a truncated M protein, composed of the N-terminal amino acid residues 1–197, with a (His)6-tag attached at the N-terminus. This recombinant protein [(His)6-Mtr], was stable but was also insoluble. After one-step affinity purification under denaturing conditions, (His)6-Mtr was used to monitor the antibody response to EMV infection by Western blot and ELISA. We obtained a 100% correlation between Western blot and virus neutralisation testing although the number of positive sera available for testing was very limited, which included seven horse, two rabbit and one human sera.

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