RESUMEN
HDM2 binds to an alpha-helical transactivation domain of p53, inhibiting its tumor suppressive functions. A miniaturized thermal denaturation assay was used to screen chemical libraries, resulting in the discovery of a novel series of benzodiazepinedione antagonists of the HDM2-p53 interaction. The X-ray crystal structure of improved antagonists bound to HDM2 reveals their alpha-helix mimetic properties. These optimized molecules increase the transcription of p53 target genes and decrease proliferation of tumor cells expressing wild-type p53.
Asunto(s)
Benzodiazepinas/síntesis química , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/agonistas , Benzodiazepinas/química , Benzodiazepinas/farmacología , Sitios de Unión , Línea Celular Tumoral , Técnicas Químicas Combinatorias , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Imitación Molecular , Estructura Molecular , Proteínas Proto-Oncogénicas c-mdm2 , Estereoisomerismo , Relación Estructura-Actividad , Proteína p53 Supresora de Tumor/biosíntesisRESUMEN
The angiotensin-converting enzyme (ACE)-related carboxypeptidase, ACE2, is a type I integral membrane protein of 805 amino acids that contains one HEXXH + E zinc-binding consensus sequence. ACE2 has been implicated in the regulation of heart function and also as a functional receptor for the coronavirus that causes the severe acute respiratory syndrome (SARS). To gain further insights into this enzyme, the first crystal structures of the native and inhibitor-bound forms of the ACE2 extracellular domains were solved to 2.2- and 3.0-A resolution, respectively. Comparison of these structures revealed a large inhibitor-dependent hinge-bending movement of one catalytic subdomain relative to the other ( approximately 16 degrees ) that brings important residues into position for catalysis. The potent inhibitor MLN-4760 ((S,S)-2-[1-carboxy-2-[3-(3,5-dichlorobenzyl)-3H-imidazol4-yl]-ethylamino]-4-methylpentanoic acid) makes key binding interactions within the active site and offers insights regarding the action of residues involved in catalysis and substrate specificity. A few active site residue substitutions in ACE2 relative to ACE appear to eliminate the S(2)' substrate-binding subsite and account for the observed reactivity change from the peptidyl dipeptidase activity of ACE to the carboxypeptidase activity of ACE2.