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A novel CXCR4-selective high-affinity fluorescent probe and its application in competitive binding assays.

ABSTRACT: We recently developed a new, rapid, and specific bioassay system that employs a fluorescent probe fabricated from our discovered CXCR4-specific ligand DV1. This new probe sensitively and selectively blocks the binding of native and synthetic ligands to CXCR4 at nanomolar levels, with a capability comparable to that seen with a conventional CXCR4 antibody. This nonradioactive, direct, and CXCR4-specific high-affinity screening system provides a new platform for CXCR4-targeted drug screening, as well as for the development of new probes for other GPCRs.

SUBMITTER: Yang Y

PROVIDER: S-EPMC4144712 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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A novel CXCR4-selective high-affinity fluorescent probe and its application in competitive binding assays.

Yang Yilei Y Zhang Qinghao Q Gao Mei M Yang Xiaohong X Huang Ziwei Z An Jing J

Biochemistry 20140728 30

We recently developed a new, rapid, and specific bioassay system that employs a fluorescent probe fabricated from our discovered CXCR4-specific ligand DV1. This new probe sensitively and selectively blocks the binding of native and synthetic ligands to CXCR4 at nanomolar levels, with a capability comparable to that seen with a conventional CXCR4 antibody. This nonradioactive, direct, and CXCR4-specific high-affinity screening system provides a new platform for CXCR4-targeted drug screening, as w ...[more]

PMID: 25058910

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Project description:CXC-type chemokine receptor 4 (CXCR4) is well known as a co-receptor for cellular entry and infection of human immunodeficiency virus type 1 (HIV-1). As an important member of the G protein-coupled receptor (GPCR) family, CXCR4 also mediates a variety of cellular processes and functions, such as cell chemotaxis, proliferation, and calcium signal transductions. Identification and characterization of molecular ligands or probes of CXCR4 have been an intensive area of investigations as such ligands or probes are of significant clinical values for the studies and treatments of HIV-1 infection and other human diseases mediated by the receptor. The crystal structures of CXCR4 in complex with different ligands have revealed two distinctive binding regions or subpockets. Thus, understanding the interactions of diverse ligands with these distinctive CXCR4 binding regions has become vital for elucidating the relationship between binding modes and biological mechanisms of ligand actions. Peptidic CVX15 is the only ligand that has been validated to bind one of these distinctive binding regions (or so called the major subpocket) of CXCR4. Therefore, in this study, we developed an efficient probe system including two high-affinity peptidic fluorescent probes, designated as FITC-CVX15 and FITC-DV1, with the aim of targeting distinctive CXCR4 subpockets. We conducted rational design and chemical characterization of the two CXCR4-specific probes and examined their application in biological experiments including competitive binding assays, flow cytometry analysis, and confocal imaging. Especially these two probes were applied in parallel CXCR4 competitive binding assays to detect and analyze potential binding modes of diverse CXCR4 ligands, together with molecular docking and simulations. Our results have indicated that these peptidic fluorescent probe systems provide novel ligand detecting tools, as well as present a new approach for analyzing distinctive binding modes of diverse CXCR4 ligands.

Dataset Information

A novel CXCR4-selective high-affinity fluorescent probe and its application in competitive binding assays.

Publications

A novel CXCR4-selective high-affinity fluorescent probe and its application in competitive binding assays.

Similar Datasets

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