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Investigation of PAS and CNBH domain interactions in hERG channels and effects of long-QT syndrome-causing mutations with surface plasmon resonance.
Soohoo, Stephanie M; Tiwari, Purushottam B; Suzuki, Yuichiro J; Brelidze, Tinatin I.
Afiliação
  • Soohoo SM; Department of Pharmacology and Physiology, Georgetown University Medical Center, Washington, District of Columbia, USA.
  • Tiwari PB; Department of Oncology, Georgetown University Medical Center, Washington, District of Columbia, USA.
  • Suzuki YJ; Department of Pharmacology and Physiology, Georgetown University Medical Center, Washington, District of Columbia, USA.
  • Brelidze TI; Department of Pharmacology and Physiology, Georgetown University Medical Center, Washington, District of Columbia, USA. Electronic address: tib5@georgetown.edu.
J Biol Chem ; 298(1): 101433, 2022 01.
Article em En | MEDLINE | ID: mdl-34801551
Human ether-á-go-go-related gene (hERG) channels are key regulators of cardiac repolarization, neuronal excitability, and tumorigenesis. hERG channels contain N-terminal Per-Arnt-Sim (PAS) and C-terminal cyclic nucleotide-binding homology (CNBH) domains with many long-QT syndrome (LQTS)-causing mutations located at the interface between these domains. Despite the importance of PAS/CNBH domain interactions, little is known about their affinity. Here, we used the surface plasmon resonance (SPR) technique to investigate interactions between isolated PAS and CNBH domains and the effects of LQTS-causing mutations R20G, N33T, and E58D, located at the PAS/CNBH domain interface, on these interactions. We determined that the affinity of the PAS/CNBH domain interactions was ∼1.4 µM. R20G and E58D mutations had little effect on the domain interaction affinity, while N33T abolished the domain interactions. Interestingly, mutations in the intrinsic ligand, a conserved stretch of amino acids occupying the beta-roll cavity in the CNBH domain, had little effect on the affinity of PAS/CNBH domain interactions. Additionally, we determined that the isolated PAS domains formed oligomers with an interaction affinity of ∼1.6 µM. Coexpression of the isolated PAS domains with the full-length hERG channels or addition of the purified PAS protein inhibited hERG currents. These PAS/PAS interactions can have important implications for hERG function in normal and pathological conditions associated with increased surface density of channels or interaction with other PAS-domain-containing proteins. Taken together, our study provides the first account of the binding affinities for wild-type and mutant hERG PAS and CNBH domains and highlights the potential functional significance of PAS/PAS domain interactions.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Síndrome do QT Longo / Proteínas Serina-Treonina Quinases / Canal de Potássio ERG1 Limite: Humans Idioma: En Revista: J Biol Chem Ano de publicação: 2022 Tipo de documento: Article País de afiliação: Estados Unidos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Síndrome do QT Longo / Proteínas Serina-Treonina Quinases / Canal de Potássio ERG1 Limite: Humans Idioma: En Revista: J Biol Chem Ano de publicação: 2022 Tipo de documento: Article País de afiliação: Estados Unidos