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Ligand Binding and Linkage

Ligand Binding and Linkage

Allosteric proteins have more than one ligand binding site; the binding of a ligand to any of these sites influences the binding of ligands to the other sites. When a protein is allosteric, its binding sites are called coupled or linked. In the case of enzymes, the site that binds to the substrate is known as the active site and the other site is known as the regulatory site. When a ligand binds to the regulatory site, this leads to conformational changes in the protein that can influence...

Ligand Binding and Linkage

Ligand Binding and Linkage

Conserved Binding Sites

Conserved Binding Sites

Many proteins’ biological role depends on their interactions with their ligands, small molecules that bind to specific locations on the protein known as ligand-binding sites. Ligand-binding sites are often conserved among homologous proteins as these sites are critical for protein function.
Binding sites are often located in large pockets, and if their location on a protein’s surface is unknown, it can be predicted using various approaches. The energetic method computationally...

Conserved Binding Sites

Conserved Binding Sites

Protein Complexes with Interchangeable Parts

Protein Complexes with Interchangeable Parts

Groups of proteins may form a complex where each protein in this complex has a different role in the overall execution of the complex’s function. Often some of the proteins in the complex can be replaced by a closely related variant to give a complex that contains many of the same components yet is functionally distinct.
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order...

Cooperative Allosteric Transitions

Cooperative Allosteric Transitions

Cooperative allosteric transitions can occur in multimeric proteins, where each subunit of the protein has its own ligand-binding site. When a ligand binds to any of these subunits, it triggers a conformational change that affects the binding sites in the other subunits; this can change the affinity of the other sites for their respective ligands. The ability of the protein to change the shape of its binding site is attributed to the presence of a mix of flexible and stable segments in the...

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Related Experiment Video

Updated: Nov 28, 2025

High Sensitivity Measurement of Transcription Factor-DNA Binding Affinities by Competitive Titration Using Fluorescence Microscopy

High Sensitivity Measurement of Transcription Factor-DNA Binding Affinities by Competitive Titration Using Fluorescence Microscopy

Published on: February 7, 2019

FGF23 contains two distinct high-affinity binding sites enabling bivalent interactions with α-Klotho.

Yoshihisa Suzuki¹, Ekaterina Kuzina¹, Seong J An¹

¹Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06510.

Proceedings of the National Academy of Sciences of the United States of America

|December 1, 2020

Summary

Fibroblast growth factor 23 (FGF23) uses two distinct repeats (R1 and R2) to bind α-Klotho (KLA), acting as a bivalent ligand. An engineered KLA-Fc fusion protein shows potential as an FGF23 antagonist.

Keywords:

biological inhibitor cell signaling endocrine FGF phosphorylation surface receptors

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Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells

Published on: April 9, 2018

Area of Science:

Endocrinology
Molecular Biology
Metabolic Regulation

Background:

Fibroblast growth factor (FGF) 23 is a key hormone regulating phosphate and vitamin D metabolism.
FGF23 signals through a complex involving α-Klotho (KLA) and FGF receptors (FGFRs).

Purpose of the Study:

To investigate the structural and functional basis of FGF23 binding to KLA.
To explore the potential of targeting the FGF23-KLA interaction for therapeutic purposes.

Main Methods:

Analysis of FGF23 C-terminal repeats (R1 and R2) for KLA binding.
Biochemical assays to measure binding affinity and receptor activation (FGFR1, MAPK).
Total internal reflection fluorescence microscopy to visualize KLA-FGFR dimerization on cell surfaces.

Main Results:

FGF23's C-terminal tail contains two functional KLA binding repeats (R1 and R2).
Both single-repeat (FGF23-R1, FGF23-R2) and wild-type FGF23 (FGF23-WT) bind KLA with similar affinity and activate signaling.
FGF23-WT functions as a bivalent ligand, promoting KLA-FGFR dimerization and activation.
Disulfide bond formation in FGF23-WT is not essential for KLA binding or signaling.

Conclusions:

FGF23 utilizes a bivalent binding mechanism with KLA through its R1 and R2 repeats.
Understanding this interaction provides insights into FGF23's metabolic regulatory functions.
An engineered KLA-Fc fusion protein acts as an FGF23 antagonist, suggesting a novel therapeutic strategy for FGF23-related disorders.