Antibody inhibition of the transcriptase activity of the rotavirus DLP: a structural view.
On entering the host cell the rotavirus virion loses its outer shell to become a double-layered particle (DLP). The DLP then transcribes the 11 segments of its dsRNA genome using its own transcriptase complex, and the mature mRNA emerges along the 5-fold axis. In order to better understand the transcription mechanism and the role of VP6 in transcription we have studied three monoclonal antibodies against VP6: RV-238 which inhibits the transcriptase activity of the DLP; and RV-133 and RV-138 which have no effect on transcription. The structures obtained by cryo-electron microscopy of the DLP/Fab complexes and by X-ray crystallography of the VP6 trimer and the VP6/Fab-238 complex have been combined to give pseudo-atomic structures. Steric hindrance between the Fabs results in limited Fab occupancy. In particular, there are on average only three of a possible five Fabs-238 which point towards the 5-fold axis. Thus, Fabs-238 are not in a position to block the exiting mRNA, nor is there any visible conformational change in VP6 on antibody binding at a resolution of 23 A. However, the epitope of the inhibiting antibody involves two VP6 monomers, whereas, those of the non-inhibiting antibodies have an epitope on only one VP6. Thus, the inhibition of transcription may be a result of inhibition of a possible change in the VP6 conformation associated with the transcription of mRNA.
Institut de Biologie Structurale, 41 rue Jules Horowitz, 38027 Grenoble, France.
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Thouvenin
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G
Schoehn
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F
Rey
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I
Petitpas
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Mathieu
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Vaney
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Cohen
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Kohli
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Pothier
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Hewat
D000914:Antibodies, Viral / Q000276:immunology*; D000956:Antigens, Viral; D002213:Capsid / Q000737:chemistry / Q000276:immunology*; D036022:Capsid Proteins; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D012321:DNA-Directed RNA Polymerases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000276:immunology; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments / Q000276:immunology / Q000494:pharmacology; D008958:Models, Molecular; D011487:Protein Conformation / Q000187:drug effects; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D012401:Rotavirus / Q000737:chemistry / Q000201:enzymology* / Q000276:immunology / Q000648:ultrastructure
D000914:Antibodies, Viral; D000956:Antigens, Viral; D036022:Capsid Proteins; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D012333:RNA, Messenger; C061332:VP6 protein, Rotavirus; D012321:DNA-Directed RNA Polymerases