In vitro reconstitution of activation of PLCepsilon by Ras and Rho GTPases.
Methods Mol Biol
; 462: 379-89, 2009.
Article
em En
| MEDLINE
| ID: mdl-19160682
ABSTRACT
Phosphatidylinositol-specific phospholipase C (PLC) enzymes catalyze the hydrolysis of phophatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] to diacylglycerol (DAG) and inositol 1,4,5-triphosphate [Ins(1,4,5)P3]. PLCepsilon is a recently discovered isoform that has been shown to be activated by members of the Ras and Rho families of guanosine trisphosphatases (GTPases) as well as subunits of heterotrimeric G-proteins. We describe a method for expressing a truncated PLCepsilon variant as an MBP fusion protein in E. coli. Subsequently, we describe the methodology necessary to reconstitute this protein with K-Ras-4B and RhoA GTPases and measure its activation.
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Base de dados:
MEDLINE
Assunto principal:
Baculoviridae
/
Proteínas ras
/
Proteínas rho de Ligação ao GTP
/
Fosfoinositídeo Fosfolipase C
Limite:
Animals
Idioma:
En
Revista:
Methods Mol Biol
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2009
Tipo de documento:
Article
País de afiliação:
Reino Unido