Use of herpes simplex virus and pseudorabies virus chimeric glycoprotein D molecules to identify regions critical for membrane fusion.
Membrane fusion induced by herpes simplex virus (HSV) requires the action of four viral membrane glycoproteins (gB, gD, gH, and gL) and the binding of gD to one of its receptors, such as the herpesvirus entry mediator or nectin-1. The related animal herpesvirus, pseudorabies virus (PRV), encodes a homologous set of glycoproteins and its gD can also use nectin-1 as an entry receptor. We show here that PRV gD, when coexpressed with HSV gB, gH, and gL, cannot substitute for HSV gD in inducing fusion with target cells expressing nectin-1. Chimeric gD molecules composed of HSV and PRV sequences can substitute, provided the first 285 aa are from HSV gD. Because the first 261 aa were sufficient for receptor binding, this suggested that amino acids 262-285 contain a region required for cell fusion but not for receptor binding. Deletions from amino acids 250-299 failed to identify a specific subregion critical for cell fusion, except possibly for amino acids 250-255, which also influenced receptor binding. Instead, presence of a flexible stalk between the membrane and receptor-binding domain appears to be required, perhaps to enable conformational changes in gD on receptor binding and subsequent interactions of undefined regions of gD with the other glycoproteins required for membrane fusion.
Zago
Anna
A
Department of Microbiology-Immunology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA.
Jogger
Cheryl R
CR
Spear
Patricia G
PG
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D015870:Gene Expression; D011558:Herpesvirus 1, Suid / Q000235:genetics* / Q000502:physiology; D008561:Membrane Fusion; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017384:Sequence Deletion / Q000235:genetics; D018139:Simplexvirus / Q000235:genetics* / Q000502:physiology; D014162:Transfection; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C039447:glycoprotein D, Human herpesvirus 1; C107752:glycoprotein D, pseudorabies virus