Video Experimental Relacionado
Updated: Aug 11, 2026
11:28
Affinity Precipitation of Active Rho-GEFs Using a GST-tagged Mutant Rho Protein (GST-RhoA(G17A)) from Epithelial Cell Lysates
Published on: March 31, 2012
Estructura del factor de intercambio guanina-nucleótido humano Mss4 e identificación de su superficie de interacción
1Howard Hughes Medical Institute, Department of Chemistry, Harvard University, Cambridge, Massachusetts 02138, USA.
Nature
|August 31, 1995
Resumen
El Mss4 humano (hMss4), un factor de intercambio de nucleótidos de guanina (GEF), tiene una estructura terciaria determinada. Esta estructura revela que su sitio activo involucra una región de unión de Zn2+ y un bucle vecino, crucial para la regulación de la proteína Rab.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Biología Estructural Biología estructural.
- Biología celular molecular Biología celular molecular
Sus antecedentes:
- Los factores de intercambio guanina-nucleótido (GEF) son reguladores clave de las Ras GTPasas, controlando el intercambio GDP/GTP y la señalización posterior.
- La rama Sec4/Ypt1/Rab de las proteínas de la superfamilia Ras juega un papel crítico en el transporte vesicular.
- El supresor de Sec4 en mamíferos (Mss4) es un EFG identificado para un subconjunto específico de proteínas Rab.
Objetivo del estudio:
- Determinar la estructura terciaria del Mss4 humano (hMss4), la primera estructura GEF de su tipo.
- Para aclarar la base estructural de la actividad del GEF de Mss4 y las interacciones de Rab-binding.
Principales métodos:
- Se empleó la espectroscopia multidimensional de resonancia magnética nuclear (RMN) para determinar la estructura terciaria de hMss4.
- Se utilizaron experimentos de perturbación de desplazamiento químico y mutagénesis dirigida al sitio para mapear la superficie de unión Rab y el sitio activo.
Principales resultados:
- Se estableció la primera estructura terciaria de una proteína GEF, hMss4, revelando una hoja beta central flanqueada por hojas más pequeñas.
- Se encontró que hMss4 se une a un ion Zn2+ a través de cuatro residuos de cisteína (Cys 23, 26, 94, 97).
- Se delinearon la superficie de unión Rab y el sitio activo, que involucra la región de unión Zn2+ y un bucle adyacente.
Conclusiones:
- La estructura determinada de hMss4 proporciona información crítica sobre el mecanismo de la actividad del GEF en la regulación de la proteína Rab.
- Comprender la interacción estructural y funcional del sitio de unión de Zn2+ y los bucles circundantes es esencial para comprender el papel de Mss4 en el transporte vesicular.
Videos de Conceptos Relacionados
02:14GTPases and their Regulation
Guanine nucleotide-binding proteins (G-proteins), also known as GTPases, are a superfamily of proteins that regulate many cellular processes, such as cell signaling, vesicular transport, and the regulation of cell shape and motility. Mutation or dysfunction of these proteins can lead to disease. There are around 40,000 known G-proteins that can broadly be classified into two groups ‒ small G-proteins consisting of a single domain and large multi-domain G-proteins.
Large G-proteins, also known...
Large G-proteins, also known...
01:33Coat Assembly and GTPases
Vesicles incorporate different coat protein subunits in different cell locations, which changes the properties of the coat, such as the shape and geometry of the transport vesicles. Thus, vesicle coat proteins also play a significant role in cargo selection.
Coat assembly depends on the local availability of phosphatidylinositol phosphates or PIPs and GTP-binding proteins. Adaptor proteins, which link the coat proteins to the membrane, bind to these PIPs and play a crucial role in controlling...
Coat assembly depends on the local availability of phosphatidylinositol phosphates or PIPs and GTP-binding proteins. Adaptor proteins, which link the coat proteins to the membrane, bind to these PIPs and play a crucial role in controlling...
01:14Rab Proteins
Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
01:25Rab Cascades
Rab GTPases act in a regulated cascade during membrane fusion, helping the lipid bilayers mix. The Rab family of proteins are active when bound to GTP, and inactive when bound to GDP. Hence, they act as guanine nucleotide-dependent molecular switches. Rab-GTP recognizes and binds to long or short-range tethering proteins to capture the target vesicle. These tethers coordinate with SNAREs on the vesicle and the target membrane to assemble the trans SNARE complex that locks the mixing bilayers.
01:24Small GTPases - Ras and Rho
Ras and Rho are small monomeric GTPases that act downstream of receptor tyrosine kinase (RTK) and regulate various cellular processes. These GTPases switch between active and inactive states by binding to guanine nucleotides.
Three regulatory proteins control their activity:
Three regulatory proteins control their activity:
01:22Activation and Inactivation of G Proteins
Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high affinity and are together...