Identification of bovine viral diarrhea virus nonstructural polypeptide NS4B/P38

The bovine pestivirus polyprotein is processed into at least 11 mature polypeptides. Previous studies with polyprotein region-specific antiserum raised against beta-galactosidase fusion proteins or synthetic peptides allowed the assignment of viral non-structural proteins to specific segments of the BVDV genome. However, the gene product from the NS4B/P38 region of the viral genome remained to be demonstrated directly. BVDV cDNA fragments predicted to encode part of NS4B/P38 (from codon 2521 to 2838 of the BVDV open reading frame) and a portion of NS5A/P58 (from codon 3008 to 3340) were expressed as glutathione S-transferase (GST) fusion proteins in Escherichia coli. Polyclonal rabbit antibodies prepared against each of the purified fusion proteins, GST-2521-2838 and GST-3008-3340, were used to immunoprecipitate viral polypeptides present in BVDV-infected cell lysates. Rabbit antiserum to GST-2521-2838 bound a polypeptide of 38 kDa identified as the mature NS4B/P38 polypeptide; while anti GST-3008-3340 lacked this specificity and bound NS5A/P58. Moreover, both antisera recognized a 96 kDa polypeptide, previously identified as a NS4B-NS5A/PP96 precursor. The function of the newly identified and highly conserved NS4B/P38 protein in viral replication remains to be determined.