Oligomerization of the human cytomegalovirus major envelope glycoprotein complex gB (gp55-116)

Abstract

The disulfide-linked glycoprotein B (gB; gp55-116) complex of human cytomegalovirus represents the most abundant and immunogenic component of the virion envelope. We have studied the oligomerization and transport of this molecule, using a series of murine monoclonal antibodies. Our results indicated that oligomerization of this molecule occurred shortly after its synthesis, with a half-time of maximal formation of approximately 25 min. The oligomeric form had an estimated mass of 340,000 Da and likely consisted of a homodimer of the gp55-116 complex. By using a conformation-specific monoclonal antibody, postoligomerization folding of this molecule was demonstrated. This event exhibited an unusually prolonged half-maximal time of approximately 160 min. Both oligomerization and folding occurred in the endoplasmic reticulum.

Cytomegalovirus-mediated activation of pyrimidine biosynthesis drives UDP–sugar synthesis to support viral protein glycosylation

“…The two cleavage products form a functional heterodimer (31,32). The different stages of gB processing are distinguishable through differential SDS/PAGE migration (31,32). We examined gB accumulation at 72 hpi in the presence of DMSO, PALA, and uridine and found that PALA treatment resulted in a decrease in the mature 116-kDa glycosylated form and an increase in the unglycosylated 105-kDa form (Fig.…”

“…The gB protein goes through several posttranslational modifications. The nascent protein (∼105 kDa) is first glycosylated, giving rise to an ∼170-kDa product, before being transported to the Golgi, where it is cleaved to form the mature protein, which consists of two fragments, one of ∼116 kDa and the other 55 kDa (31,32). The two cleavage products form a functional heterodimer (31,32).…”

“…The nascent protein (∼105 kDa) is first glycosylated, giving rise to an ∼170-kDa product, before being transported to the Golgi, where it is cleaved to form the mature protein, which consists of two fragments, one of ∼116 kDa and the other 55 kDa (31,32). The two cleavage products form a functional heterodimer (31,32). The different stages of gB processing are distinguishable through differential SDS/PAGE migration (31,32).…”

Development of novel vaccines against human cytomegalovirus

“…[71][72][73] However, in studies to express recombinant HCMV gB, the inclusion of the furin cleavage site led to low yields of monomeric gB, whereas the elimination of this site by mutation resulted in efficient production, but synthesis of mostly monomeric gB, with some higher MW forms, using a variety of mammalian and insect cells. [74][75][76][77] Recently, mutations to the fusion loops of a HCMV gB consisting of amino acid residues 78-706 resulted in a trimeric gB produced in insect cells, with the structure subsequently analyzed by X-ray crystallography. 78 Another trimeric HCMV gB in a postfusion conformation was produced and consisted of amino acid residues 86-698 bound to the Fab fragments of a neutralizing human anti-gB antibody, with the structure also analyzed by X-ray crystallography.…”

The Human Cytomegalovirus-Specific UL1 Gene Encodes a Late-Phase Glycoprotein Incorporated in the Virion Envelope

“…Many of the HCMV structural glycoproteins form high-molecular-weight disulfide-linked oligomers, such as the gCI com- plex, composed of multidimers of gB (9,39); therefore, we analyzed UL1 protein migration patterns under both nonreducing and reducing (with ␤-mercaptoethanol) conditions. The UL1 protein was ␤-mercaptoethanol resistant, implying that it is not part of a disulfide-linked oligomer in infected MRC-5 cells (data not shown).…”