Effects of JZTX-V on the wild type Kv4.3 Expressed in HEK293T and Molecular Determinants in the Voltage-sensing Domains of Kv4.3 Interacting with JZTX-V

Xu, Donghui; Wenmei, Wu; Siqin, Hong; Zeng, Peng; Wang, Xianchun; Zeng, Xin

doi:10.1080/19336950.2022.2053420

Cited by 4 publications

(1 citation statement)

References 34 publications

(47 reference statements)

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“…The site-directed mutagenesis confirms that the three residues S301 in the S4 and Y312, L321 in the channel S4-S5 linker are important for gliquidone-mediated Kv4.3 inhibition. These findings are consistent with the observations that the four-helical bundle voltage sensing S1-S4 domains (VSDs) and the S4-S5 linker of voltage gated channels are important pharmacological targets for their binding with small molecules and peptide toxins (Abbott, 2020;Ahuja et al, 2015;Dehong et al, 2022;DeSimone et al, 2009;Montero-Domínguez et al, 2022;Xu et al, 2019).…”

Section: Discussionsupporting

confidence: 91%

Inhibition of Cardiac Kv4.3/KChIP2 Channels by Sulfonylurea Drug Gliquidone

Yang,

Li,

et al. 2023

Mol Pharmacol

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The Kv4.3 channel is featured of fast N-type inactivation and also undergoes a slow C-type inactivation. The gain-of-function mutations of Kv4.3 channels cause an inherited disease called Brugada syndrome characterized by a shortened duration of cardiac action potential repolarization and ventricular arrhythmia. The sulfonylurea drug gliquidone, an ATP-dependent K + channel antagonist, is widely used for the treatment of type 2 diabetes.Here, we report a novel role of gliquidone in inhibiting Kv4.3 and Kv4.3/KChIP2 channels that encode the cardiac Ito currents responsible for the initial phase of action potential repolarization. Gliquidone results in concentration-dependent inhibition of both Kv4.3 and Kv4.3/KChIP2 fast or steady-state inactivation currents with an IC 50 of approximately 8 μM.Gliquidone also accelerates Kv4.3 channel inactivation and shifts the steady-state activation to a more depolarizing direction. Site-directed mutagenesis and molecular docking reveal that the residues S301 in the S4 and Y312A, L321A in the S4-S5 linker are critical for gliquidone-mediated inhibition of Kv4.3 currents, as mutating those residues to alanine significantly reduces the potency for gliquidone-mediated inhibition. Furthermore, gliquidone also inhibits a gain-of-function Kv4.3 V392I mutant identified in BrS patients in voltage-and concentration-dependent manner. Taken together, our findings demonstrate that gliquidone inhibits Kv4.3 channels by acting on the residues in the S4 and the S4-S5 linker. Therefore, gliquidone may hold repurposing potential for the therapy of Brugada syndrome. Significance StatementWe describe a novel role of gliquidone in inhibiting cardiac Kv4.3 currents and the channel gain-of-function mutation identified from patients with Brugada syndrome, suggesting its repurposing potential for therapy for the heart disease.

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Section: Discussionsupporting

confidence: 91%

Inhibition of Cardiac Kv4.3/KChIP2 Channels by Sulfonylurea Drug Gliquidone

Yang,

Li,

et al. 2023

Mol Pharmacol

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Structure–Function Relationship and Stability of Latroeggtoxin‐VI: A Proteinaceous Toxin From the Eggs of Latrodectus tredecimguttatus

Chen,

Sun,

Yin

et al. 2024

Peptide Science

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Latroeggtoxin‐VI (LETX‐VI), a peptide toxin discovered from the eggs of spider Latrodectus tredecimguttatus, was previously shown to promote the synthesis and release of dopamine in rat pheochromocytoma (PC12) cells, showing potential applications in the neurobiology and medicine. To further understand the structure and properties of LETX‐VI, the key residues were identified and their roles in the structure, function, and stability of LETX‐VI were analyzed in the present work. Based on the protein molecular docking, our previous work, and the relevant literature, the potential key residues of LETX‐VI were selected and identified by alanine‐scanning mutagenesis. The wild‐type LETX‐VI and its 13 mutants, including a double mutant, were prepared by gene cloning and heterologous expression in Escherichia coli, followed by activity, structure, and stability determination. The results demonstrated that the activity of the mutants K25A, R35A, K40A/R41A, and L45A, particularly R35A, to promote dopamine release from PC12 cells was significantly decreased compared with that of the wild‐type LETX‐VI, indicating that these mutated residues are the key residues. Circular dichroism (CD) analysis showed that the secondary structure of these mutants was not obviously different from that of wild‐type LETX‐VI, suggesting that mutation‐caused decrease in the activity of LETX‐VI is due to the changes in the binding site on the molecule surface, rather than the abnormal alternation of the molecular conformation of LETX‐VI. Acetonitrile (ACN) did not obviously influence the activity of LETX‐VI; however, 0.1% trifluoroacetic acid (TFA) treatment for 2 h significantly reduced its activity. Treatment with weakly acidic and basic buffers (pH ≥ 6.6) for 12 h was favorable for LETX‐VI and R35A to exert their activity. Higher temperatures (>37°C) decreased the activity of both wild‐type LETX‐VI and R35A. In conclusion, K25, R35, K40, R41, and L45 particularly R35 are the important functional site residues; during experiments, care should be taken to avoid the adverse influence of strong acid and high temperature on LETX‐VI. These observations have enhanced our understanding of the structure and properties of LETX‐VI and provided references for the subsequent modification of structure and function of LETX‐VI.

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