The complete nucleotide sequence of the gammaherpesvirus equine herpesvirus 5 (EHV5) glycoprotein B (gB) was determined and the deduced amino acid sequence compared with that of the second equine gammaherpesvirus EHV2. EHV5 gB is an 870 amino acid protein and is 79% similar and 66% identical with EHV2 gB at the amino acid level. EHV5 gB like EHV2 gB is a disulphide linked heterodimer with subunits of 92 and 68 kDa. EHV5 gB is an integral membrane glycoprotein containing only N-linked oligosaccharides and contains a putative endoproteolytic cleavages site at amino acids 422-485. The EHV5 gB amino acid sequence showed greatest homology with other members of the Rhadinovirus genus of the subfamily Gammaherpesvirinae. Alignment of EHV5 gB sequence with the gB sequence of seven other gammaherpesviruses showed conservation of 10 cysteine residues as well as conservation of three predicted sites of N-linked glycosylation; the highest degree of conservation of the predicted sites of N-linked glycosylation was observed between EHV5 and the other members of the Rhadinovirus genus. Phylogenetic analysis confirmed EHV2 and EHV5 were most closely related to each other and equally distant from other members of the Rhadinovirus genus included in the analysis.