Scholarlink

Abstract <p>The intermediate-conductance calcium-activated potassium channel K<sub>Ca</sub>3.1 promotes calcium-dependent hyperpolarization of the cell membrane. Its malfunction has been observed in autoimmune and oncological diseases. To study this channel and its peptide blockers using fluorescence analysis, plasmids encoding the α-subunit K<sub>Ca</sub>3.1 fused with the fluorescent protein mKate2 at the N- or C-terminus were constructed, and the fluorescent ligand ChTx-GFP was obtained, which is a combination of the peptide blocker charybdotoxin and the green fluorescent protein. It was found that mKate2 at the N-terminus of the α-subunit blocks the transport of the channel into the plasma membrane of Neuro-2a cells, while mKate2 at its C-terminus does not interfere with the efficient accumulation of the channel in the plasma membrane and the formation of a regular tetrameric structure capable of binding peptide blockers. The ligand ChTx-GFP binds to the K<sub>Ca</sub>3.1 channel on the membrane at a concentration of 20 nM and can be used for fluorescent imaging of these channels in mammalian cells.</p>

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

N- or C-terminal Position of the Fluorescent Protein mKate2 in the mKate2-KCa3.1 Chimera Influences Membrane Expression of the Channel

  • 期刊论文

  • 发表日期：

    2025年12月1日

查看全文
  • V. N. Korabeynikova,
  • A. V. Feofanov,
  • O. V. Nekrasova

摘要

Abstract

The intermediate-conductance calcium-activated potassium channel KCa3.1 promotes calcium-dependent hyperpolarization of the cell membrane. Its malfunction has been observed in autoimmune and oncological diseases. To study this channel and its peptide blockers using fluorescence analysis, plasmids encoding the α-subunit KCa3.1 fused with the fluorescent protein mKate2 at the N- or C-terminus were constructed, and the fluorescent ligand ChTx-GFP was obtained, which is a combination of the peptide blocker charybdotoxin and the green fluorescent protein. It was found that mKate2 at the N-terminus of the α-subunit blocks the transport of the channel into the plasma membrane of Neuro-2a cells, while mKate2 at its C-terminus does not interfere with the efficient accumulation of the channel in the plasma membrane and the formation of a regular tetrameric structure capable of binding peptide blockers. The ligand ChTx-GFP binds to the KCa3.1 channel on the membrane at a concentration of 20 nM and can be used for fluorescent imaging of these channels in mammalian cells.