Structure of the regulatory N-domain of human cardiac troponin C in complex with human cardiac troponin I147-163 and bepridil

Xu Wang, Monica X. Li, Brian D. Sykes

Research output: Contribution to journalArticle

62 Scopus citations

Abstract

Cardiac troponin C (cTnC) is the Ca2+-dependent switch for contraction in heart muscle and a potential target for drugs in the therapy of heart failure. Ca2+ binding to the regulatory domain of cTnC (cNTnC) induces little structural change but sets the stage for cTnI binding. A large "closed" to "open" conformational transition occurs in the regulatory domain upon binding ctnI147-163 or bepridil. This raises the question of whether cTnI147-163 and bepridil compete for cNTnC·Ca2+. In this work, we used two-dimensional 1H,15N-heteronuclear single quantum coherence (HSQC) NMR spectroscopy to examine the binding of bepridil to cNTnC·Ca2+ in the absence and presence of cTnI147-163 and of ctnI147-163 to cNTnC·Ca2+ in the absence and presence of bepridil. The results show that bepridil and ctnI147-163 bind cNTnC·Ca2+ simultaneously but with negative cooperativity. The affinity of cTnI147-163 for cNTnC·Ca2+ is reduced ∼3.5-fold by bepridil and vice versa. Using multinuclear and multidimensional NMR spectroscopy, we have determined the structure of the cNTnC· Ca2+·cTnI147-163·bepridil ternary complex. The structure reveals a binding site for cTnI147-163 primarily located on the A/B interhelical interface and a binding site for bepridil in the hydrophobic pocket of cNTnC·Ca2+. In the structure, the N terminus of the peptide clashes with part of the bepridil molecule, which explains the negative cooperativity between cTnI147-163 and bepridil for cNTnC·Ca2+. This structure provides insights into the features that are important for the design of cTnC-specific cardiotonic drugs, which may be used to modulate the Ca2+ sensitivity of the myofilaments in heart muscle contraction.

Original languageEnglish (US)
Pages (from-to)31124-31133
Number of pages10
JournalJournal of Biological Chemistry
Volume277
Issue number34
DOIs
StatePublished - Aug 23 2002
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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