RESUMEN
The highly coordinated interactions of several molecular chaperones, including hsp70 and hsp90, are required for the folding and conformational regulation of a variety of proteins in eukaryotic cells, such as steroid hormone receptors and many other signal transduction regulators. The protein called Hop serves as an adaptor protein for hsp70 and hsp90 and is thought to optimize their functional cooperation. Here we characterize the assembly of the hsp70-Hop-hsp90 complex and reveal interactions that cause conformational changes between the proteins in the complex. We found that hsp40 plays an integral role in the assembly by enhancing the binding of hsp70 to the Hop complex. This is accomplished by stimulating the conversion of hsp70-ATP to hsp70-ADP, the hsp70 conformation favored for Hop binding. The hsp70-Hop-hsp90 complex is highly dynamic, as has been observed previously for hsp90 in its interaction with client proteins. Nonetheless, hsp90 binds with high affinity to Hop (K(d) = 90 nm), and this binding is not affected by hsp70. hsp70 binds with lower affinity to Hop (K(d) = 1.3 microm) on its own, but this affinity is increased (K(d) = 250 nm) in the presence of hsp90. hsp90 also reduces the number of hsp70 binding sites on the Hop dimer from two sites in the absence of hsp90 to one site in its presence. Hop can inhibit the ATP binding and p23 binding activity of hsp90, yet this can be reversed if hsp70 is present in the complex. Taken together, our results suggest that the assembly of hsp70-Hop-hsp90 complexes is selective and influences the conformational state of each protein.
Asunto(s)
Proteínas HSP70 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Chaperonas Moleculares/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Sistema Libre de Células , Proteínas del Choque Térmico HSP40 , Proteínas HSP70 de Choque Térmico/genética , Proteínas HSP90 de Choque Térmico/genética , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Sustancias Macromoleculares , Chaperonas Moleculares/genética , Unión ProteicaRESUMEN
The progesterone receptor (PR) can be isolated in its native conformation able to bind hormone, yet its ligand-binding domain rapidly loses its activity at elevated temperature. However, an in vitro chaperoning system consisting of five proteins (HSP40, HSP70, HOP, HSP90, and p23) with ATP is capable of restoring this function. The first step of this chaperoning mechanism is usually thought to be the binding of HSP70 to PR. Our findings here show that the binding of HSP40 to PR is, instead, the first step. HSP40 binding occurred rapidly and was not dependent on ATP or other proteins. The stoichiometry of HSP40 to native PR in these complexes was approximately 1:1. HSP40 bound specifically and with a high affinity to native PR (K(d) = 77 nm). The binding of HSP40 to PR was sustained and did not interact in the highly dynamic fashion that has been observed previously for HSP90 in this system. The HSP40 small middle dotPR complex could be isolated as a functional unit that could, after the addition of the other chaperones, progress to a PR complex capable of hormone binding. These results indicate that HSP40 initiates the entry of PR into the HSP90 pathway.
Asunto(s)
Proteínas de Choque Térmico/metabolismo , Receptores de Progesterona/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Pollos , ADN Complementario/metabolismo , Electroforesis en Gel de Poliacrilamida , Proteínas del Choque Térmico HSP40 , Proteínas HSP70 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Oxidorreductasas Intramoleculares , Cinética , Ratones , Modelos Químicos , Chaperonas Moleculares/metabolismo , Fosfoproteínas/metabolismo , Prostaglandina-E Sintasas , Unión Proteica , Pliegue de Proteína , Isoformas de Proteínas , Temperatura , Factores de TiempoRESUMEN
Una forma de identificación Médico Legal es la rconstitución facial en osamentas humanas. Un problema que se presenta es la determinación de la forma, tamaño y ubicación de estructuras del rostro, que no presentan, al parecer, relación directa con las estructuras craneanas, como por ejemplo la nariz, labios y ojos. Es sabido, que la proporción natural en los seres vivientes es la proporción aurea, y el ser humano mantiene esta relación de proporcionalidad en las dimensiones de las distintas estructuras que conforman su cuerpo. Matemáticamente esta proporción es la relación de la unidad (1) con 1,618 veces la misma (1,618=Ý). El presente trabajo se realizó con un universo de 40 individuos de ambos sexos, entre 21 y 30 años, de los que se obtuvieron las dimensiones de diferentes estructuras del cráneo, por medio de telerradiografías y de distintas estructuras del rostro. por medio de diapositivas, que tenían relación de proporcionalidad áurea entre ellas. Los resultados entregaron un total de 355 relaciones de proporcionalidad áurea cráneo-facial, craneanas y faciales