Tripartite split-GFP assay to identify selective intracellular nanobody that suppresses GTPase RHOA subfamily downstream signaling.

ABSTRACT: Strategies based on intracellular expression of artificial binding domains present several advantages over manipulating nucleic acid expression or the use of small molecule inhibitors. Intracellularly-functional nanobodies can be considered as promising macrodrugs to study key signaling pathways by interfering with protein-protein interactions. With the aim of studying the RAS-related small GTPase RHOA family, we previously isolated, from a synthetic phage display library, nanobodies selective towards the GTP-bound conformation of RHOA subfamily proteins that lack selectivity between the highly conserved RHOA-like and RAC subfamilies of GTPases. To identify RHOA/ROCK pathway inhibitory intracellular nanobodies, we implemented a stringent, subtractive phage display selection towards RHOA-GT

SUBMITTER: Keller L 

PROVIDER: S-EPMC9433928 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature