Posttranslational folding of vesicular stomatitis virus G protein in the ER: involvement of noncovalent and covalent complexes.
In this study, we show that posttranslational folding of Vesicular Stomatitis virus G protein subunits can involve noncovalent, multimeric complexes as transient intermediates. The complexes are heterogeneous in size (4-21S20,W), contain several G glycopolypeptides, and are associated with BiP/GRP78. The newly synthesized, partially intrachain disulfide-bonded G proteins enter these complexes immediately after chain termination, and are released 1-4 min later as fully oxidized, trimerization-competent monomers. These monomers are properly folded, judging by their binding of conformation-specific mAbs. When the G protein is translated in the presence of DTT, it remains reduced, largely unfolded and aggregated in the ER, but it can fold successfully when the DTT is removed. In this case, contrary to normal folding, the aggregates become transiently disulfide cross-linked. We also demonstrated that the fidelity of the folding process is dependent on metabolic energy. Finally, we established that the G protein of the folding mutant of the Vesicular Stomatitis virus, ts045, is blocked at a relatively late step in the folding pathway and remains associated with oligomeric, BiP/GRP78-containing folding complexes.
Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut 06510-8002.
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de Silva
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Braakman
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Helenius
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D016466:CHO Cells; D002499:Centrifugation, Density Gradient; D006224:Cricetinae; D004229:Dithiothreitol / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D004734:Energy Metabolism; D008562:Membrane Glycoproteins; D008954:Models, Biological; D014176:Protein Biosynthesis; D011487:Protein Conformation; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics* / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D014759:Viral Envelope Proteins; D000255:Adenosine Triphosphate; D004229:Dithiothreitol