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STD and TRNOESY NMR studies for the epitope mapping of the phosphorylation motif of the oncogenic protein beta-catenin recognized by a selective monoclonal antibody.

Megy S, Bertho G, Gharbi-Benarous J, Baleux F, Benarous R, Girault JP
FEBS Lett 580 5411-22 (2006)
Cited: 5 times
EuropePMC logo PMID: 16996060

Abstract

The interaction of the P-beta-Cat(19-44) peptide, a 26 amino acid peptide (K(19)AAVSHWQQQSYLDpSGIHpSGATTTAP(44)) that mimics the phosphorylated beta-Catenin antigen, has been studied with its monoclonal antibody BC-22, by transferred nuclear Overhauser effect NMR spectroscopy (TRNOESY) and saturation transfer difference NMR (STD NMR) spectroscopy. This antibody is specific to diphosphorylated beta-Catenin and does not react with the non-phosphorylated protein. Phosphorylation of beta-Catenin at sites Ser33 and Ser37 on the DSGXXS motif is required for the interaction of beta-Catenin with the ubiquitin ligase SCF(beta-TrCP). beta-TrCP is involved in the ubiquitination and proteasome targeting of the oncogenic protein beta-Catenin, the accumulation of which has been implicated in various human cancers. The three-dimensional structure of the P-beta-Cat(19-44) in the bound conformation was determined by TRNOESY NMR experiments; the peptide adopts a compact structure in the presence of mAb with formation of turns around Trp25 and Gln26, with a tight bend created by the DpS(33)GIHpS(37) motif; the peptide residues (D32-pS37) forming this bend are recognized by the antibody as demonstrated by STD NMR experiments. STD NMR studies provide evidence for the existence of a conformational epitope containing tandem repeats of phosphoserine motifs. The peptide's epitope is predominantly located in the large bend and in the N-terminal segment, implicating bidentate association. These findings are in excellent agreement with a recently published NMR structure required for the interaction of beta-Catenin with the SCF(beta-TrCP) protein.

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  2. Molecular dynamic simulation of mutated β-catenin in solid pseudopapillary neoplasia of the pancreas. Tipmanee V, Pattaranggoon NC, Kanjanapradit K, Saetang J, Sangkhathat S. Oncol Lett 15 9167-9173 (2018)


Articles citing this publication (3)

  1. GSK-3 phosphorylates delta-catenin and negatively regulates its stability via ubiquitination/proteosome-mediated proteolysis. Oh M, Kim H, Yang I, Park JH, Cong WT, Baek MC, Bareiss S, Ki H, Lu Q, No J, Kwon I, Choi JK, Kim K. J Biol Chem 284 28579-28589 (2009)
  2. Epitope mapping using ribosome display in a reconstituted cell-free protein synthesis system. Osada E, Shimizu Y, Akbar BK, Kanamori T, Ueda T. J Biochem 145 693-700 (2009)
  3. Characterization of the prostaglandin H2 mimic: binding to the purified human thromboxane A2 receptor in solution. Ruan KH, Wijaya C, Cervantes V, Wu J. Arch Biochem Biophys 477 396-403 (2008)


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