Referenced in: none

Automatically associated channels: KChIP1 , Kir2.3 , Kv1.4 , Kv3.1 , Kv4.3

Title: Interactions of KChIP4a and its mutants with Ca2+ or Kv4.3 N-terminus by affinity capillary electrophoresis.

Authors: Meina Li, Lei Lei, Linghan Jia, Xiaomei Ling, Jianmei Zhang, Yiran Zhao, Kewei Wang

Journal, date & volume: Anal. Biochem., 2014 Mar 15 , 449, 99-105

PubMed link: http://www.ncbi.nlm.nih.gov/pubmed/24361715

Abstract
The specific binding of auxiliary Kv channel-interacting proteins (KChIPs) to the N terminus of Kv4 pore-forming α-subunits results in modulation of gating properties, surface expression, and subunit assembly of Kv4 channels. However, the interactions between KChIPs and Kv4 remain elusive. Thus, affinity capillary electrophoresis (ACE) was employed to quantitatively evaluate the interactions between KChIPs and Kv4.3 N terminus (KvN) and between KChIP4a/related mutants and Ca(2+) for the first time. The mobility ratio, derivatives calculated from the mobility shift method, was used to deduce the binding constants (Kb). As a result, the binding constants for KChIP4a/KvN and KChIP1/KvN complexes were (8.32±1.66)×10(6) L mol(-1) and (5.26±0.71)×10(6) L mol(-1), respectively. In addition, in the presence of calcium (10 μmol L(-1)), the binding constant of KChIP4a/KvN increased to (6.72±1.66)×10(7) L mol(-1). In addition, the binding constant of KChIP4a with Ca(2+) was (7.1±1.5)×10(7) L mol(-1). Besides, studies on the effect of truncated mutants revealed that the third EF hand of KChIP4a was related to high-affinity binding with Ca(2+), and the integrity of the molecular structure of KChIP4a was important for Ca(2+) binding. This method profits from small samples, rapid analysis, and simple operation without being time-consuming.