RESUMO
Inhibition of the ATPase activity of the chaperone protein HSP90 is a potential strategy for treatment of cancers. We have determined structures of the HSP90alpha N-terminal domain complexed with the purine-based inhibitor, PU3, and analogs with enhanced potency both in enzyme and cell-based assays. The compounds induce upregulation of HSP70 and downregulation of the known HSP90 client proteins Raf-1, CDK4, and ErbB2, confirming that the molecules inhibit cell growth by a mechanism dependent on HSP90 inhibition. We have also determined the first structure of the N-terminal domain of HSP90beta, complexed with PU3. The structures allow a detailed rationale to be developed for the observed affinity of the PU3 class of compounds for HSP90 and also provide a structural framework for design of compounds with improved binding affinity and drug-like properties.
Assuntos
Adenina/análogos & derivados , Adenina/química , Adenosina Trifosfatases/antagonistas & inibidores , Anisóis/química , Inibidores Enzimáticos/química , Proteínas de Choque Térmico HSP90/química , Purinas/química , Adenina/metabolismo , Adenina/farmacologia , Anisóis/metabolismo , Anisóis/farmacologia , Sítios de Ligação , Divisão Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Humanos , Ligantes , Modelos Moleculares , Estrutura Molecular , Isoformas de Proteínas , Estrutura Terciária de Proteína , Purinas/metabolismo , Purinas/farmacologia , Relação Estrutura-AtividadeRESUMO
Urease allows organisms to use exogenous and internally generated urea as a nitrogen source, by catalyzing the hydrolysis of urea to form ammonia and carbon dioxide. Urease may also participate in the systemic nitrogen-transport pathways and possibly acts as a toxic defence protein. Jack bean urease (JBU) was the first nickel-metalloenzyme identified and was crystallized as early as 1926. Despite this, the structure has not yet been determined. An antibody fragment, Fv, that has a high affinity for JBU has been used to aid crystallization. The complex, which retains full enzyme activity, forms very small crystals that diffract weakly to 3.3 A. The crystals belong to the rhombohedral space group R32, with unit-cell parameters a = b = 228.6, c = 130.9 A. The structure of the urease molecule has been solved by molecular replacement using the structure of homogenous enzyme from Klebsiella aerogenes as a search model.