Structural models of the bovine papillomavirus E5 protein

Surti, Toral; Klein, Ophir D.; Aschheim, Kathryn; DiMaio, Daniel; Smith, Steven O.

doi:10.1002/(sici)1097-0134(19981201)33:4<601::aid-prot12>3.0.co;2-i

Cited by 46 publications

(65 citation statements)

References 45 publications

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“…It contains a hydrophobic central segment and exists in the intracellular membranes of transformed cells as a type II TM homodimer stabilized by disulfide bonds in its carboxy-terminus (3)(4)(5). The E5 dimer binds to the TM domains of two molecules of the platelet-derived growthfactor β receptor (PDGFβR) tyrosine kinase, resulting in receptor dimerization and activation, leading to cell transformation (6)(7)(8)(9)(10).…”

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confidence: 99%

Construction and genetic selection of small transmembrane proteins that activate the human erythropoietin receptor

Cammett¹,

Jun²,

Cohen³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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This work describes a genetic approach to isolate small, artificial transmembrane (TM) proteins with biological activity. The bovine papillomavirus E5 protein is a dimeric, 44-amino acid TM protein that transforms cells by specifically binding and activating the platelet-derived growth factor β receptor (PDGFβR). We used the E5 protein as a scaffold to construct a retrovirus library expressing ∼500; 000 unique 44-amino acid proteins with randomized TM domains. We screened this library to select small, dimeric TM proteins that were structurally unrelated to erythropoietin (EPO), but specifically activated the human EPO receptor (hEPOR). These proteins did not activate the murine EPOR or the PDGFβR. Genetic studies with one of these activators suggested that it interacted with the TM domain of the hEPOR. Furthermore, this TM activator supported erythroid differentiation of primary human hematopoietic progenitor cells in vitro in the absence of EPO. Thus, we have changed the specificity of a protein so that it no longer recognizes its natural target but, instead, modulates an entirely different protein. This represents a novel strategy to isolate small artificial proteins that affect diverse membrane proteins. We suggest the word "traptamer" for these transmembrane aptamers.bovine papilloma virus | E5 protein | expression libraries | protein engineering | traptamers I t is thought that up to 30% of all proteins span cell membranes (1). Most membrane-spanning protein segments adopt an α-helical conformation that is largely independent of their amino acid sequence and minimally influenced by extramembraneous protein domains. Although transmembrane (TM) domains were once thought to be rather nonspecific protein segments whose primary function was to anchor proteins in membranes, they can participate in highly-specific, inter-and intramolecular side-by-side interactions that control important biological processes (2).The 44-amino acid bovine papillomavirus type 1 (BPV) E5 protein is essentially an isolated TM domain. It contains a hydrophobic central segment and exists in the intracellular membranes of transformed cells as a type II TM homodimer stabilized by disulfide bonds in its carboxy-terminus (3-5). The E5 dimer binds to the TM domains of two molecules of the platelet-derived growthfactor β receptor (PDGFβR) tyrosine kinase, resulting in receptor dimerization and activation, leading to cell transformation (6-10). The E5 protein does not bind to other TM domains or to certain PDGFβR TM mutants (11).Based on our analysis of the E5 protein, we proposed that it might be possible to construct small, artificial TM proteins that regulate a variety of viral and cellular TM proteins in trans (11, 12). To identify small proteins with different TM sequences that transform cells, we constructed expression libraries in which the TM domain of the E5 protein was replaced with randomized sequences of primarily hydrophobic amino acids and isolated clones from these libraries that induced focus formation or growth-factor independe...

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mentioning

confidence: 99%

Construction and genetic selection of small transmembrane proteins that activate the human erythropoietin receptor

Cammett¹,

Jun²,

Cohen³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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“…On the basis of these studies and computational analysis, several models of the complex between the E5 protein and the PDGFβR TMD have been proposed (4,6,9,12,23). In each of these models, the complex consists of a single dimer of the E5 protein bridging two molecules of the PDGFβR TMD, but the stoichiometry of the complex has not been established.…”

Section: Significancementioning

confidence: 99%

“…The E5 protein is an extremely hydrophobic integral membrane protein located primarily in the membranes of the Golgi apparatus of transformed cells (2,3). Biophysical studies in model membranes indicate that the E5 protein adopts a transmembrane orientation roughly perpendicular to the membrane surface (4)(5)(6). In essence, the E5 protein is a freestanding transmembrane domain (TMD), with a type II transmembrane orientation in which a short C-terminal segment protrudes into the lumen of the Golgi (2).…”

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confidence: 99%

Two transmembrane dimers of the bovine papillomavirus E5 oncoprotein clamp the PDGF β receptor in an active dimeric conformation

Karabadzhak

Petti

Barrera

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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The dimeric 44-residue E5 protein of bovine papillomavirus is the smallest known naturally occurring oncoprotein. This transmembrane protein binds to the transmembrane domain (TMD) of the platelet-derived growth factor β receptor (PDGFβR), causing dimerization and activation of the receptor. Here, we use Rosetta membrane modeling and all-atom molecular dynamics simulations in a membrane environment to develop a chemically detailed model of the E5 protein/PDGFβR complex. In this model, an active dimer of the PDGFβR TMD is sandwiched between two dimers of the E5 protein. Biochemical experiments showed that the major PDGFβR TMD complex in mouse cells contains two E5 dimers and that binding the PDGFβR TMD to the E5 protein is necessary and sufficient to recruit both E5 dimers into the complex. These results demonstrate how E5 binding induces receptor dimerization and define a molecular mechanism of receptor activation based on specific interactions between TMDs.transmembrane protein complex | oncogene | traptamer | BPV | blue native gel electrophoresis B ecause viruses modulate signaling nodes that control cell behavior and virus replication, the study of viral proteins has provided great insight into many aspects of cellular biochemistry and the cellular processes that regulate biological function. Thus, viral proteins have long been recognized as valuable tools to probe central problems in biology. A particularly interesting viral protein is the 44-residue E5 oncoprotein encoded by bovine papillomavirus type 1 (BPV). The BPV E5 protein and closely related E5 proteins of other fibropapillomaviruses are the shortest known, naturally occurring proteins with tumorigenic potential (1). The E5 protein is an extremely hydrophobic integral membrane protein located primarily in the membranes of the Golgi apparatus of transformed cells (2, 3). Biophysical studies in model membranes indicate that the E5 protein adopts a transmembrane orientation roughly perpendicular to the membrane surface (4-6). In essence, the E5 protein is a freestanding transmembrane domain (TMD), with a type II transmembrane orientation in which a short C-terminal segment protrudes into the lumen of the Golgi (2). In cells, detergent micelles, and model lipid bilayers, the E5 protein exists as a homodimer stabilized by disulfide bonds involving C-terminal cysteine residues (3-5, 7-10). Genetic studies showed that E5 dimerization is required for transforming activity, and a preferred orientation of the E5 dimer with a symmetric homodimer interface has been identified (7,9,11,12).The E5 protein transforms cells by activating the cellular platelet-derived growth factor (PDGF) β receptor (PDGFβR), although there may be additional minor, alternative transforming pathways as well (13). The PDGFβR is a receptor tyrosine kinase (RTK) with an extracellular domain that binds PDGF, a hydrophobic membrane-spanning segment, and a cytoplasmic domain with tyrosine kinase activity. The inactive PDGFβR is primarily monomeric in unstimulated cells, and PDGF binding indu...

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“…The E5 protein is the major transforming protein of BPV in cultured fibroblasts (DiMaio et al, 1986;Schiller et al, 1986). In transformed cells, the E5 protein exists as a disulfidelinked type II transmembrane homodimer localized largely to the membranes of the endoplasmic reticulum and Golgi apparatus (Schlegel et al, 1986;Burkhardt et al, 1987Burkhardt et al, , 1989Surti et al, 1998). Transformation requires Gln17, Asp33, and two conserved cysteine residues in the carboxy-terminus that mediate disulfide bond formation, but activity is not impaired by a variety of hydrophobic substitution mutations in the central hydrophobic domain of the E5 protein (Horwitz et al, 1988;Meyer et al, 1994).…”

Section: The Importance Of Transmembrane Interactionsmentioning

confidence: 99%

Modulation of cell function by small transmembrane proteins modeled on the bovine papillomavirus E5 protein

Freeman-Cook

DiMaio

2005

Oncogene

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Structural models of the bovine papillomavirus E5 protein

Cited by 46 publications

References 45 publications

Construction and genetic selection of small transmembrane proteins that activate the human erythropoietin receptor

Construction and genetic selection of small transmembrane proteins that activate the human erythropoietin receptor

Two transmembrane dimers of the bovine papillomavirus E5 oncoprotein clamp the PDGF β receptor in an active dimeric conformation

Modulation of cell function by small transmembrane proteins modeled on the bovine papillomavirus E5 protein

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