Modification and activation of Ras proteins by electrophilic prostanoids with different structure are site-selective.
Cyclopentenone prostanoids (cyP) arise as important modulators of inflammation and cell proliferation. Although their physiological significance has not been fully elucidated, their potent biological effects have spurred their study as leads for the development of therapeutic agents. A key determinant of cyP action is their ability to bind to thiol groups in proteins or in glutathione through Michael addition. Even though several protein targets for cyP addition have been identified, little is known about the structural determinants from the protein or the cyP that drive this modification. The results herein presented provide the first evidence that cyP with different structures target distinct thiol sites in a protein molecule, namely, H-Ras. Whereas 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) and Delta12-PGJ2 preferentially target the C-terminal region containing cysteines 181 and 184, PGA1 and 8-iso-PGA1 bind mainly to cysteine 118, located in the GTP-binding motif. The biological counterparts of this specificity are the site-selective modification and activation of H-Ras in cells and the differential interaction of cyP with H, N, and K-Ras proteins. Cysteine 184 is unique to H-Ras, whereas cysteine 118 is present in the three Ras homologues. Consistent with this, PGA1 binds to and activates H-, N-, and K-Ras, thus differing from the preferential interaction of 15d-PGJ2 with H-Ras. These results put forward the possibility of influencing the selectivity of cyP-protein addition by modifying cyP structure. Furthermore, they may open new avenues for the development of cyP-based drugs.
Renedo
Marta
M
Departamento de Estructura y Función de Proteínas, Centro de Investigaciones Biológicas, C.S.I.C., Ramiro de Maeztu 9, 28040 Madrid, Spain.
Gayarre
Javier
J
García-Domínguez
Carlota A
CA
Pérez-Rodríguez
Andrea
A
Prieto
Alicia
A
Cañada
F Javier
FJ
Rojas
José M
JM
Pérez-Sala
Dolores
D
D000818:Animals; D000893:Anti-Inflammatory Agents / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites / Q000187:drug effects; D003545:Cysteine / Q000737:chemistry / Q000187:drug effects; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D015230:Prostaglandin D2 / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D011453:Prostaglandins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D011454:Prostaglandins A / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry; D014407:Tumor Cells, Cultured; D018631:ras Proteins / Q000737:chemistry / Q000145:classification / Q000187:drug effects* / Q000378:metabolism*
D000893:Anti-Inflammatory Agents; D011453:Prostaglandins; D011454:Prostaglandins A; D011994:Recombinant Proteins; C499810:delta(12)-prostaglandin J(2); C037112:9-deoxy-delta-9-prostaglandin D2; D018631:ras Proteins; D003545:Cysteine; D015230:Prostaglandin D2; C100573:prostaglandin A1