Involvement of the molecular chaperone BiP in maturation of Sindbis virus envelope glycoproteins.
Sindbis virus codes for two membrane glycoproteins, E1 and PE2, which assemble into heterodimers within the endoplasmic reticulum. We have examined the role of the molecular chaperone BiP (grp78) in the maturation of these two proteins. E1, which folds into its mature conformation via at least three intermediates differing in the configurations of their disulfide bonds, was found to interact strongly and transiently with BiP after synthesis. ATP depletion mediated by carbonyl cyanide m-chlorophenylhydrazone treatment results in the stabilization of complexes between BiP and E1. The depletion of intracellular ATP levels also greatly inhibits conversions between the E1 folding intermediates and results in the slow incorporation of E1 into disulfide-stabilized aggregates. These results suggest that the ATP-regulated binding and release of BiP have a role in modulating disulfide bond formation during E1 folding. In comparison with E1, very little PE2 is normally recovered in association with BiP. However, under conditions in which E1 folding is aberrant, increased amounts of PE2 become directly associated with BiP. The formation of these BiP-PE2 interactions occurs after E1 begins to misfold or fails to fold efficiently. We propose that nascent PE2 is stable prior to pairing with E1 for only a limited period of time, after which unpaired PE2 becomes recognized by BiP. This implies that the productive association of PE2 and E1 must occur within a restricted time frame and only after E1 has accomplished certain folding steps mediated by BiP binding and release. Kinetic studies which show that the pairing of E1 with PE2 is delayed after translocation support this conclusion.
Cell Research Institute, University of Texas at Austin 78713-7640.
M
M
Mulvey
D T
DT
Brown
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D006224:Cricetinae; D004220:Disulfides; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism*; D011485:Protein Binding; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D012845:Sindbis Virus / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000378:metabolism*
D002352:Carrier Proteins; D004220:Disulfides; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins; D018832:Molecular Chaperones; D014759:Viral Envelope Proteins; C022012:glycoprotein E1, Sindbis virus; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D000255:Adenosine Triphosphate