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J Virol. 2010 Feb;84(4):1696-703. doi: 10.1128/JVI.01968-09. Epub 2009 Dec 2.

Membrane orientation of the human papillomavirus type 16 E5 oncoprotein.

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1
Department of Pathology, Georgetown University Medical School, 3900 Reservoir Rd. NW, Washington, DC 20057, USA.

Abstract

The E5 protein of human papillomavirus type 16 is a small, hydrophobic protein that localizes predominantly to membranes of the endoplasmic reticulum (ER). To define the orientation of E5 in these membranes, we employed a differential, detergent permeabilization technique that makes use of the ability of low concentrations of digitonin to selectively permeabilize the plasma membrane and saponin to permeabilize all cellular membranes. We then generated a biologically active E5 protein that was epitope tagged at both its N and C termini and determined the accessibility of these termini to antibodies in the presence and absence of detergents. In both COS cells and human ectocervical cells, the C terminus of E5 was exposed to the cytoplasm, whereas the N terminus was restricted to the lumen of the ER. Finally, the deletion of the E5 third transmembrane domain (and terminal hydrophilic amino acids) resulted in a protein with its C terminus in the ER lumen. Taken together, these topology findings are compatible with a model of E5 being a 3-pass transmembrane protein and with studies demonstrating its C terminus interacting with cytoplasmic proteins.

PMID:
19955310
PMCID:
PMC2812368
DOI:
10.1128/JVI.01968-09
[Indexed for MEDLINE]
Free PMC Article
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