Vaccinia virus has two infectious forms, the intracellular mature virus, which has a single envelope, and the extracellular enveloped virus (EEV), which is surrounded by two lipid bilayers. The outer membrane of the EEV contains at least six viral proteins. Among them A34, a type II membrane glycoprotein, and B5, a type I membrane glycoprotein, form a complex and are involved in processes such as morphogenesis and EEV entry. A34 is required for normal incorporation of B5 into the EEV membrane. Here, we used a virus lacking B5 and viruses with mutations in the B5 membrane-proximal stalk region and looked at the effect of those modifications on A34. Data presented show that B5 is required for the correct glycosylation, trafficking and stability of A34, emphasizing the complex interactions and mutual dependence of these vaccinia EEV proteins.