IDENTIFICATION OF AN INFECTIOUS LARYNGOTRACHEITIS VIRUS GENE ENCODINGAN IMMUNOGENIC PROTEIN WITH A PREDICTED M(R) OF 32 KILODALTONS

The nucleotide sequence of an infectious laryngotracheitis virus (ILTV ) gene which maps immediately upstream from the glycoprotein 60 (gp60) gene was determined. The gene, designated p32, encodes a predicted po lYPeptide of 298 amino acids with an estimated M(r) of 32000 daltons. The predicted protein sequence has four potential N-glycosylation site s and a signal sequence at the N-terminal region. Amino acid residues in the NH2-terminal region of the p32 protein exhibit similarity to gl ycoprotein X (gX) of pseudorabies virus (PRV) and its homolog in equin e herpesvirus type 1 (EHV-1). Within the conserved (N-terminus) region , one putative N-linked glycosylation site and four cysteine residues are aligned in these proteins. These common structural features of the gX-like proteins were also found in glycoprotein G (gG) of human herp es simplex virus type 2 (HSV-2) and equine herpesvirus type 4 (EHV-4). High level bacterial production of the p32 protein was achieved by cl oning the p32 open reading frame into a pGEX-2T expression vector. Wes tern blot analysis of the fusion protein produced in E. coli using imm une chicken sera confirms that p32 protein is of viral origin and is a n immunogen in birds with infectious laryngotracheitis (ILT). An antis erum from chicken immunized with the fusion protein detected a substan tial amount of p32 protein in the medium of ILTV-infected cells in Wes tern blotting. Moreover tunicamycin treatment of cells infected with t he virus indicated that p32 was glycosylated. This allows us to conclu de that p32 is a glycoprotein and like gX of PRV accumulates in the me dium of infected cells.