ISOLATION OF A TRIPEPTIDE FROM A RANDOM PHAGE PEPTIDE LIBRARY THAT INHIBITS P-1,P-4-DIADENOSINE 5'-TETRAPHOSPHATE BINDING TO ITS RECEPTOR

Extracellular P-1,P-4-diadenosine 5'-tetraphosphate (Ap(4)A) has been implicated as a modulator of cell stress. We have previously demonstra ted specific receptors for Ap(4)A at the surface of cardiac myocytes ( Walker et al., 1993a). In addition, we have isolated a monoclonal anti body (mAb TL4) that recognized the Ap(4)A receptor and inhibited bindi ng of Ap(4)A to its receptor (Walker & Hilderman, 1993). As part of ou r effort to characterize the Ap(4)A receptor building domain, we scree ned a random phage peptide library with mAb TL4. After affinity purifi cation of specifically bound phage, we isolated 38 individual phage cl ones. Twenty-eight of these clones bound mAb TL4 in ELISA and dot blot analyses. Twenty-two of the twenty-eight individual clones contained inserts with an RGS tripeptide sequence. Synthetic RGS peptide specifi cally inhibits the binding of mAb TL4 to its membrane receptor. Furthe rmore, the RGS peptide also inhibits [H-3]Ap(4)A binding to its recept or. These data are consistent with the RGS peptide mimicking part of t he mAb TL4 recognition site on the Ap(4)A receptor. The RGS peptide ma y be used to help characterize the Ap(4)A receptor binding domain and to help determine the physiological significance of the interaction be tween Ap(4)A and its receptor.