TR-FRET between engineered nanobodies reveals the existence of endogenous CXCR4 oligomers.

ABSTRACT: Chemokine receptors CXCR4 and ACKR3 are involved in multiple physiological and pathological processes. In overexpression systems, CXCR4 and ACKR3 form oligomers that are important for chemokine recognition and signaling. Whether this holds true in physiological settings is unknown as no evidence of such oligomers are available. To address this gap of knowledge, we develop a new approach for GPCR oligomer detection by time-resolved Förster resonance energy transfer (TR-FRET) between fluorescently labeled nanobodies. Fluorescent probes are conjugated to the C-terminus of extracellularly binding nanobodies using site-directed sortase-mediated labeling, retaining high affinity of the nanobodies for their target receptors. This approach, first validated in transfected cells by detecting the pre

SUBMITTER: Heuninck J 

PROVIDER: S-EPMC12708788 | biostudies-literature | 2025 Dec

REPOSITORIES: biostudies-literature