title: Expression of bacteriorhodopsin in Sf9 and COS-1 cells.
abstract: We report studies on the expression of the archaebacterial membrane protein bacteriorhodopsin in Sf9 insect cells and in COS-1 mammalian cells. In both cell systems, the apoprotein bacterio-opsin was expressed at levels of approximately 1 microgram/10(6) cells. Immunofluorescence studies showed that the expressed protein was accumulated in the endoplasmic reticulum. However, upon addition of all-trans retinal to membranes isolated from either Sf9 or COS-1 cells expressing bacterio-opsin, the characteristic bacteriorhodopsin chromophore (lambda max at approximately 560 nm) was rapidly generated. This is in contrast to bacterio-opsin expressed in E. coli, which cannot be functionally reconstituted with retinal unless it is first denatured, and then renatured in vitro. These studies demonstrate that the bacterio-opsin expressed is correctly folded and show that localization of a heterologously expressed membrane protein in the endoplasmic reticulum does not necessarily imply that it is misfolded.
authors.affiliation: Department of Biological Chemistry, Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA.
authors.forename: J
authors.initials: J
authors.lastname: Heymann
authors.forename: R
authors.initials: R
authors.lastname: Jager
authors.forename: S
authors.initials: S
authors.lastname: Subramaniam
mesh_terms: D000818:Animals; D001436:Bacteriorhodopsins / Q000235:genetics* / Q000378:metabolism; D019556:COS Cells; D002460:Cell Line; D004721:Endoplasmic Reticulum; D015870:Gene Expression; D051379:Mice; D017510:Protein Folding; D018411:Spodoptera / Q000166:cytology
chemical_list: D001436:Bacteriorhodopsins