Moloney murine leukemia virus envelope protein subunits, gp70 and Pr15E, form a stable disulfide-linked complex.
The nature and stability of the interactions between the gp70 and Pr15E/p15E molecules of murine leukemia virus (MLV) have been disputed extensively. To resolve this controversy, we have performed quantitative biochemical analyses on gp70-Pr15E complexes formed after independent expression of the amphotropic and ecotropic Moloney MLV env genes in BHK-21 cells. We found that all cell-associated gp70 molecules are disulfide linked to Pr15E whereas only a small amount of free gp70 is released by the cells. The complexes were resistant to treatment with reducing agents in vivo, indicating that the presence and stability of the disulfide interaction between gp70 and Pr15E are not dependent on the cellular redox state. However, disulfide-bonded Env complexes were disrupted in lysates of nonalkylated cells in a time-, temperature-, and pH-dependent fashion. Disruption seemed not to be caused by a cellular factor but is probably due to a thiol-disulfide exchange reaction occurring within the Env complex after solubilization. The possibility that alkylating agents induce the formation of the intersubunit disulfide linkage was excluded by showing that disulfide-linked gp70-Pr15E complexes exist in freshly made lysates of nonalkylated cells and that disruption of the complexes can be prevented by lowering the pH. Together, these data establish that gp70 and Pr15E form a stable disulfide-linked complex in vivo.
Opstelten
D J
DJ
Department for Biosciences at Novum, Karolinska Institute, S-141 57 Huddinge, Sweden. Dirk-Jan.Opstelten@cbt.ki.se
Wallin
M
M
Garoff
H
H
D000478:Alkylation; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D004220:Disulfides / Q000378:metabolism*; D004229:Dithiothreitol / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008979:Moloney murine leukemia virus / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D019163:Reducing Agents / Q000494:pharmacology; D015682:Retroviridae Proteins, Oncogenic / Q000378:metabolism*; D012995:Solubility; D013696:Temperature; D013997:Time Factors; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*
D004220:Disulfides; D019163:Reducing Agents; D015682:Retroviridae Proteins, Oncogenic; D014759:Viral Envelope Proteins; C078678:p15E protein, Murine leukemia virus; D004229:Dithiothreitol