Related Experiment Video
Updated: Jul 19, 2026

Expression and Purification of Mammalian Bestrophin Ion Channels
Published on: August 2, 2018
The bestrophin mutation A243V, linked to adult-onset vitelliform macular dystrophy, impairs its chloride channel
Kuai Yu1, Yuanyuan Cui, H Criss Hartzell
1Department of Cell Biology, The Center for Neurodegenerative Disease, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322, USA.
Purpose:
It has been proposed that Best vitelliform macular dystrophy (BVMD) is caused by dysfunction in the Cl(-) channel function of human bestrophin-1 (hBest1), but some patients with BVMD who have the hBest1 A243V mutation have normal electro-oculograms, suggesting that this mutation may not affect Cl(-) channel function. The purpose of this study was to determine whether the A243V mutation affects the Cl(-) channel function of hBest1.
Methods:
Wild-type alanine at position 243 was changed to valine by PCR-based mutagenesis. Wild-type (WT) and A243V hBest1 were transfected into HEK-293 cells, and Cl(-) currents were measured with the whole-cell patch-clamp technique. The trafficking of proteins to the plasma membrane was tested by cell-surface biotinylation.
Results:
WT hBest1 induced Ca(2+)-activated Cl(-) currents in HEK cells that were >1 nA in amplitude. The currents produced by the A243V mutant, however, were only approximately 10% as large as WT. This was not due to the inability of the A243V mutant to reach the plasma membrane, as shown by cell-surface biotinylation. The A243V mutation changed channel anion selectivity. The WT current exhibited a relative permeability P(X)/P(Cl) order of SCN(-) > or = I(-) > or = NO(3)(-) > Br(-) > Cl(-) > HCO(3)(-) and a relative conductance G(X)/G(Cl) order of NO(3)(-) > SCN(-) > I(-) > or = Br(-) > or = Cl(-) > HCO(3)(-). However, the A243V current exhibited different sequences: P(X)/P(Cl) was SCN(-) > NO(3)(-) > I(-) > Br(-) > Cl(-) > HCO(3)(-) and G(X)/G(Cl) was SCN(-) > NO(3)(-) > or = I(-) > or = Br(-) > Cl(-) > HCO(3)(-). Unlike several other hBest1 mutations that have dominant-negative effects on wild-type channels, the A243V-mutation did not influence the wild-type current when A243V and WT hBest1 were transfected together.
Conclusions:
The disease-causing A243V mutation is associated with altered hBest1 Cl(-) channel activity. The absence of a dominant negative effect of A243V is consistent with the more mild symptoms associated with this mutation. These results are interpreted in terms of the hypotheses that bestrophins are Cl(-) channels and regulators of Ca signaling.
Related Concept Videos
Cystic Fibrosis: Pathogenesis
CF is primarily caused by a genetic mutation in a chromosome 7 gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein. The most common gene mutation leading to CF is the ΔF508 mutation, but...
Translation
Translation is the process of synthesizing proteins from the genetic information carried by messenger RNA (mRNA). Following transcription, it constitutes the final step in the expression of genes. This process is carried out by ribosomes, complexes of protein and specialized RNA molecules. Ribosomes, transfer RNA (tRNA), and other proteins produce a chain of amino acids—the polypeptide—as the end product of translation.
Translation Produces the Building Blocks of Life
Translation
Translation is the process of synthesizing proteins from the genetic information carried by messenger RNA (mRNA). Following transcription, it constitutes the final step in the expression of genes. This process is carried out by ribosomes, complexes of protein and specialized RNA molecules. Ribosomes, transfer RNA (tRNA), and other proteins produce a chain of amino acids—the polypeptide—as the end product of translation.
Translation Produces the Building Blocks of Life
Sex-linked Disorders
Lysosomal Hydrolases
ATP Synthase: Mechanism
