Transcriptional and structural analyses of Amsacta moorei entomopoxvirus protein kinase gene (AMV197, pk)

Abstract

The Amsacta moorei entomopoxvirus (AMEV) genome has 279 open reading frames (ORFs) among which is the AMV197, composed of 900 nt and potentially encoding a protein of 299 amino acids. Sequence-derived amino acid analysis suggested it to be a serine/threonine protein kinase (PK) having conserved PK and serine/threonine PK domains. For transcriptional analysis of the AMV197 pk gene, Ld652 cells were infected with AMEV and mRNA was isolated at different times thereafter. RT–PCR analysis indicated that the transcription of the AMV197 pk gene started at 4 h post infection (h p.i.) and continued to be expressed through 24 h p.i. Infection of Ld cells in the presence of Ara-C (inhibits DNA replication), followed by RT–PCR showed that AMV197 pk is transcribed as an early gene. Transcription was initiated at 54 nt upstream of the translation start site. The vaccinia virus early promoter element G was also found at the correct position (−21) in the AMV197 pk gene. Rapid amplification of the 3′ ends of the AMV197 pk transcript showed that there are two polyadenylation start points. They are located at 22 and 32 nucleotides downstream of translation stop site. Also, the translational stop site and poly (A) signal of AMV197 pk are overlapped. The termination signal TTTTTGT sequence of vaccinia virus early genes was found just upstream of the 3′ end of AMV197 pk gene. Conserved amino acid subdomains of the AMV197 PK were found by sequence comparisons with PK’s from other organisms. Analysis of the protein sequence of AMV197 pk gene reveals close identity with PK genes of other organisms.