A bacterial multi-hybrid system and applications

Details A bacterial multi-hybrid system and applications A bacterial multi-hybrid system and applications

G - Physics

01

N

G01N 33/50 (2006.01) C12Q 1/02 (2006.01) C12Q 1/527 (2006.01) C12Q 1/68 (2006.01) G01N 33/542 (2006.01) G01N 33/58 (2006.01)

Patent

CA 2312379

A signal amplification system comprises a bacterial multi-hybrid system, and more preferably a two-hybrid system, of at least two chimeric polypeptides containing a first chimeric polypeptide corresponding to a first fragment of an enzyme and a second chimeric polypeptide corresponding to a second fragment of an enzyme or a modulating substance capable of activating said enzyme. The first fragment is fused to a molecule of interest and the second fragment or the modulating substance is fused to a target ligand. The activity of the enzyme is restored by the in vivo interaction between the molecule of interest and the target ligand. Signal amplification is generated and, for example, triggers transcriptional activation. The signal amplification system is useful in a method of selecting a molecule of interest, which is capable of binding to target ligand, wherein the interaction between the molecule of interest and the target ligand is detected with the signal amplification system as a kit therefor. A method of screening for a substance capable of stimulating or inhibiting the interaction between a target ligand and a molecule of interest is also provided.

Système d'amplification de signal consistant en un système bactérien comportant des hybrides multiples et, de préférence, deux hybrides, et composé d'au moins deux polypeptides chimères contenant un premier polypeptide chimère correspondant à un premier fragment d'une enzyme et un deuxième polypeptide chimère correspondant à un deuxième fragment d'une enzyme ou à une substance de modulation capable d'activer ladite enzyme. On fusionne ce premier fragment avec une molécule ciblée et on fusionne le deuxième fragment ou la substance de modulation avec un ligand ciblé. On rétablit l'activité de l'enzyme par l'interaction in vivo entre la molécule et le ligand ciblés. On génère une amplification de signal, ce qui déclenche, par exemple, l'activation de la transcription. Ce système d'amplification de signal est utile dans un procédé servant à sélectionner une molécule capable de se fixer au ligand ciblé, l'interaction entre cette molécule et le ligand ciblé étant détectée par ce système sous forme de trousse. L'invention concerne également un procédé de recherche par criblage d'une substance capable de stimuler ou d'inhiber l'interaction entre un ligand ciblé et une molécule.

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