Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Related Concept Videos

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...

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 Sites

Ligand Binding Sites

Proteins are dynamic macromolecules that carry out a wide variety of essential processes; however, the activities of most proteins depend on their interactions with other molecules or ions, known as ligands.
Protein-ligand interactions are quite specific; even though numerous potential ligands surround a cellular protein at any given time, only a particular ligand can bind to that protein. Moreover, a ligand binds only to a dedicated area on the surface of the protein, known as the...

Allosteric Proteins-ATCase

Allosteric Proteins-ATCase

Binding sites linkages can regulate a protein's function. For example, enzyme activity is often regulated through a feedback mechanism where the end product of the biochemical process serves as an inhibitor.
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis...

Non-gated Ion Channels

Non-gated Ion Channels

Ion channels are specialized proteins on the plasma membrane that allow charged ions to pass down their electrochemical gradient. Their main function is to maintain the membrane potential which is critical for cell viability. These channels are either gated or non-gated and can transport more than a thousand ions within milliseconds for the cellular event to occur.
Compared to the gated ion channels, the non-gated channels, also known as leakage or passive channels, have no gating mechanism....

Ligand-Gated Ion Channel Receptor: Gating Mechanism

Ligand-Gated Ion Channel Receptor: Gating Mechanism

Ligand-gated ion channels are transmembrane proteins that play a vital role in intercellular communication and functions of the nervous system. They allow the influx of ions across the membrane once the neurotransmitter binds, allowing the subsequent transmission of electrical excitation across the neurons. Other ligand-gated ion channels, like the γ-aminobutyric acid (GABA) receptor, permit anions like chloride into the cells on the binding of the GABA molecule. Their entry into the cell...

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by

Same author

Serological evidence of SARS-CoV-2 exposure in marine mammals in the United States between 2020 and 2025.

PloS one·2026

Same author

Severe neonatal epilepsy linked to a novel homozygous SLC12A5 Arg508Cys variant with impaired chloride transport.

Experimental neurology·2026

Same author

Exploring sedentary behaviour reduction interventions in the primary healthcare setting: a scoping review.

BMC primary care·2026

Same author

Compound heterozygous SLC12A5 variants expand the molecular and functional spectrum of KCC2-developmental and epileptic encephalopathy.

Epilepsia·2026

Same author

Retinal degeneration protein 3 mutants are associated with cell-cycle arrest and apoptosis.

Cell death discovery·2025

Same author

Circulating tumor cell plasticity determines breast cancer therapy resistance via neuregulin 1-HER3 signaling.

Nature cancer·2025

Search research articles

Related Experiment Video

Updated: May 13, 2025

Study of the Functions and Activities of Neuronal K-Cl Co-Transporter KCC2 Using Western Blotting

Study of the Functions and Activities of Neuronal K-Cl Co-Transporter KCC2 Using Western Blotting

Published on: December 9, 2022

Highly conserved ion binding sites are not all functionally relevant in mouse KCC4.

Lisa Becker¹, Jens Hausmann², Rieke Wellpott¹

¹Division of Neurogenetics, School of Medicine and Health Science, Carl von Ossietzky University of Oldenburg, Oldenburg, Germany.

Frontiers in Molecular Biosciences

|April 15, 2025

Summary

Potassium chloride cotransporter 4 (KCC4) shows flexibility in ion coordination sites, unlike KCC2. Specific residues are crucial for potassium and chloride binding, with implications for transporter function.

Keywords:

KCC ion binding sites large extracellular loop protein conformation site directed mutagenesis

More Related Videos

Mutagenesis and Functional Analysis of Ion Channels Heterologously Expressed in Mammalian Cells

Mutagenesis and Functional Analysis of Ion Channels Heterologously Expressed in Mammalian Cells

Published on: October 1, 2010

Isolation and Kv Channel Recordings in Murine Atrial and Ventricular Cardiomyocytes

Isolation and Kv Channel Recordings in Murine Atrial and Ventricular Cardiomyocytes

Published on: March 12, 2013

Related Experiment Videos

Last Updated: May 13, 2025

Study of the Functions and Activities of Neuronal K-Cl Co-Transporter KCC2 Using Western Blotting

Study of the Functions and Activities of Neuronal K-Cl Co-Transporter KCC2 Using Western Blotting

Published on: December 9, 2022

Mutagenesis and Functional Analysis of Ion Channels Heterologously Expressed in Mammalian Cells

Mutagenesis and Functional Analysis of Ion Channels Heterologously Expressed in Mammalian Cells

Published on: October 1, 2010

Isolation and Kv Channel Recordings in Murine Atrial and Ventricular Cardiomyocytes

Isolation and Kv Channel Recordings in Murine Atrial and Ventricular Cardiomyocytes

Published on: March 12, 2013

Area of Science:

Biochemistry
Molecular Biology
Structural Biology

Background:

Potassium chloride cotransporter 4 (KCC4) is vital for kidney acidification and hearing.
KCCs are known to transport potassium (K+) and chloride (Cl-) ions.
Cryo-electron microscopy (CryoEM) suggested two distinct chloride (Cl-) binding sites in KCC4.

Purpose of the Study:

To investigate the functional importance of residues coordinating potassium and chloride ions in KCC4.
To compare ion coordination requirements between KCC4 and KCC2.

Main Methods:

Point mutations were introduced to residues involved in ion coordination in KCC4.
Thallium (Tl+) based flux measurements were employed to assess transporter activity.

Main Results:

Not all conserved coordination sites in KCC4 are essential for function.
Specific residues were identified as critical for potassium coordination (N131, Y216, T432).
Residues for chloride coordination in Cl1 (G134, V135, I136) and Cl2 (G433, M435, Y589) were identified, showing differential importance compared to KCC2.

Conclusions:

KCC4 exhibits greater flexibility in ion coordination stringency than KCC2.
The large extracellular loop (LEL) and other structural elements likely contribute to these differences.