Detection of intracellular enzyme complex

Details Detection of intracellular enzyme complex Detection of intracellular enzyme complex

C - Chemistry, Metallurgy

12

N

C12N 15/87 (2006.01) C12N 15/00 (2006.01)

Patent

CA 2627022

Methods and compositions are provided for determining intracellular translocation of proteins employing .beta.-galactosidase fragments that independently complex to form an active enzyme. Cells are used having two fusion constructs: one fragment bound to a protein of interest; and the other fragment bound to a compartment localizing signal. The cells are used to screen compounds for their effect on translocation, where a substrate containing high ionic strength solution is used for detection of the enzyme complex.

L'invention concerne des méthodes et des compositions pour déterminer une translocation intracellulaire de protéines au moyen de fragments de ß-galactosidase complexant indépendamment pour former une enzyme active. Selon l'invention, des cellules utilisées ont deux structures de fusion, à savoir un fragment lié à une protéine d'intérêt et l'autre fragment lié à un signal de localisation de compartiment. Les cellules servent au criblage de composés pour vérifier leur effet sur la translocation, un substrat contenant une solution de force ionique élevée servant à la détection du complexe enzymatique.

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