Sequence-based design of a peptide probe for the APC tumor suppressor protein

Sharma, Victoria A.; Logan, Jennifer S.; King, David S.; White, Ray; Alber, Tom

doi:10.1016/s0960-9822(98)70324-0

Cited by 33 publications

(35 citation statements)

References 54 publications

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“…In the absence of additional experiments, one potential concern is that some of the associations we find might occur nonspecifically (i.e., are false positives) (25). Three lines of evidence, however, suggest that many of the interactions reported here are specific.…”

Section: Resultsmentioning

confidence: 84%

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A computationally directed screen identifying interacting coiled coils from Saccharomyces cerevisiae

Newman

Wolf

Kim

2000

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

152

134

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Computational methods can frequently identify protein-interaction motifs in otherwise uncharacterized open reading frames. However, the identification of candidate ligands for these motifs (e.g., so that partnering can be determined experimentally in a directed manner) is often beyond the scope of current computational capabilities. One exception is provided by the coiled-coil interaction motif, which consists of two or more ␣ helices that wrap around each other: the ligands for coiled-coil sequences are generally other coiled-coil sequences, thereby greatly simplifying the motif͞ligand recognition problem. Here, we describe a twostep approach to identifying protein-protein interactions mediated by two-stranded coiled coils that occur in Saccharomyces cerevisiae. Coiled coils from the yeast genome are first predicted computationally, by using the MULTICOIL program, and associations between coiled coils are then determined experimentally by using the yeast two-hybrid assay. We report 213 unique interactions between 162 putative coiled-coil sequences. We evaluate the resulting interactions, focusing on associations identified between components of the spindle pole body (the yeast centrosome).

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Section: Resultsmentioning

confidence: 84%

“…Fourth, coiled coils are often capable of mediating specific interactions without the need for additional protein domains (20,24,25). Fifth, coiled coils occur in a large number and wide range of proteins of general interest, including structural proteins, motor proteins, transcription factors, and membrane fusion proteins (9,12,26,27).…”

mentioning

confidence: 99%

A computationally directed screen identifying interacting coiled coils from Saccharomyces cerevisiae

Newman

Wolf

Kim

2000

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

152

134

View full text Add to dashboard Cite

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“…The principle behind this has been exploited previously to generate reagents that block the function of the coiled coil containing proteins APC and human immunodeficiency virus gp41 (4,15,29,37) and is illustrated in Fig. 1A.…”

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confidence: 99%

The Zipper Region of Epstein-Barr Virus bZIP Transcription Factor Zta Is Necessary but Not Sufficient To Direct DNA Binding

Hicks

Al-Mehairi

Sinclair

2003

J Virol

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The viral bZIP transcription factor Zta (BZLF1, EB1, ZEBRA) mediates the switch between the latent and lytic cycles of Epstein-Barr virus (EBV). In part, its activity requires the formation of homodimers and interaction with specific DNA sequence elements (ZREs). Zta has an atypical zipper motif that has a lower stability than do typical bZIP proteins. Here we show that a synthetic peptide directed against the zipper can disrupt the DNA-binding function of Zta. This highlights the relevance of this region for the function of Zta and demonstrates that the zipper region is a potential target for therapeutic agents. We also unmask the relevance of a region adjacent to the zipper (CT region), which is required to direct the interaction of Zta with DNA and to transactivate ZRE-dependent promoters in vivo.

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“…First, the N-terminal 250 amino acids form the stretch with the highest portion of structural elements. This domain includes the Nterminal 55 amino acids, which form a coiled coil region that is responsible for the dimerization of the APC protein (22)(23)(24). The N-terminal 250 amino acids also include the two leucinerich motifs, residues 68 -77 and 165-174, which were proposed as NES motifs (12,15).…”

Section: Resultsmentioning

confidence: 99%

“…At this stage of our study, there were only preliminary structural data available for this region (22,23). We purified the recombinantly expressed protein fragment APC-(2-250) and found that APC-(2-250) forms oligomeric complexes (Fig.…”

Section: Resultsmentioning

confidence: 99%

The Coiled Coil Region (Amino Acids 129–250) of the Tumor Suppressor Protein Adenomatous Polyposis Coli (APC)

Tickenbrock

Cramer

Vetter

et al. 2002

Journal of Biological Chemistry

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The APC (adenomatous polyposis coli) tumor suppressor protein has many different intracellular functions including a nuclear export activity. Only little is known about the molecular architecture of the 2843-amino acid APC protein. Guided by secondary structure predictions we identified a fragment close to the N-terminal end, termed APC-(129 -250), as a soluble and proteaseresistant domain. We solved the crystal structure of APC-(129 -250), which is monomeric and consists of three ␣-helices forming two separate antiparallel coiled coils. APC-(129 -250) includes the nuclear export signal NES-(165-174) at the C-terminal end of the first helix. Surprisingly, the conserved hydrophobic amino acids of NES-(165-174) are buried in one of the coiled coils and are thus not accessible for interaction with other proteins. We demonstrate the direct interaction of APC-(129 -250) with the nuclear export factor chromosome maintenance region 1 (Crm-1). This interaction is enhanced by the small GTPase Ran in its activated GTPbound form and also by a double mutation in APC-(129 -250), which deletes two amino acids forming two of the major interhelical interactions within the coiled coil. These observations hint to a regulatory mechanism of the APC nuclear export activity by NES masking.The tumor suppressor gene APC 1 is inactivated in the majority of human colorectal cancers. One major function of the APC protein is the regulation of the level and intracellular localization of the signaling proto-oncoprotein ␤-catenin (1). Together with other proteins the APC protein increases the turnover rate of cytosolic ␤-catenin and thereby decreases the intracellular ␤-catenin level (2-5). Stabilizing mutations in ␤-catenin or truncating mutations in APC lead to an increase of the intracellular ␤-catenin concentration (6 -8). As a consequence, ␤-catenin can enter the nucleus where it activates transcription in complex with a transcription factor of the T-cell transcription factor/lymphoid enhancer binding factor family (9, 10). In addition to regulating the cytosolic ␤-catenin level, the APC protein can also regulate the intracellular localization of ␤-catenin and its nuclear signaling activity. The APC protein has a nuclear export function and may shuttle ␤-catenin between the nucleus and the cytoplasm (1). The nuclear accumulation of APC, induced by the inhibition of the exportin Crm-1 (chromosome maintenance region 1), leads to the accumulation of ␤-catenin in the nucleus (12-14). The APC protein contains several leucine-rich motifs, which might function as pertinent nuclear export signals (NESs), suggesting that APC interacts directly with Crm-1 (15). Two leucine-rich motifs, which were proposed as NES motifs, are located at the N terminus including residues 68 -77 and 165-174, respectively (12, 14, 15). MATERIALS AND METHODSSubcloning, Mutagenesis, Expression, and Protein Purification-The full-length APC cDNA was kindly provided by Paul Polakis (Genentech). The fragments APC-(2-250) and APC-(129 -250) were PCR-amplified. The double muta...

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Sequence-based design of a peptide probe for the APC tumor suppressor protein

Cited by 33 publications

References 54 publications

A computationally directed screen identifying interacting coiled coils from Saccharomyces cerevisiae

A computationally directed screen identifying interacting coiled coils from Saccharomyces cerevisiae

The Zipper Region of Epstein-Barr Virus bZIP Transcription Factor Zta Is Necessary but Not Sufficient To Direct DNA Binding

The Coiled Coil Region (Amino Acids 129–250) of the Tumor Suppressor Protein Adenomatous Polyposis Coli (APC)

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