Structural and Functional
Consequences of the Cardiac
Troponin C L48Q Ca<sup>2+</sup>-Sensitizing Mutation
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Abstract
Calcium binding to the regulatory domain of cardiac troponin
C
(cNTnC) causes a conformational change that exposes a hydrophobic
surface to which troponin I (cTnI) binds, prompting a series of protein–protein
interactions that culminate in muscle contraction. A number of cTnC
variants that alter the Ca<sup>2+</sup> sensitivity of the thin filament
have been linked to disease. Tikunova and Davis engineered a series
of cNTnC mutations that altered Ca<sup>2+</sup> binding properties
and studied the effects on the Ca<sup>2+</sup> sensitivity of the
thin filament and contraction [Tikunova, S. B., and Davis, J. P. (2004) <i>J. Biol. Chem. 279</i>, 35341–35352]. One of the mutations
they engineered, the L48Q variant, resulted in a pronounced increase
in the cNTnC Ca<sup>2+</sup> binding affinity and Ca<sup>2+</sup> sensitivity
of cardiac muscle force development. In this work, we sought structural
and mechanistic explanations for the increased Ca<sup>2+</sup> sensitivity
of contraction for the L48Q cNTnC variant, using an array of biophysical
techniques. We found that the L48Q mutation enhanced binding of both
Ca<sup>2+</sup> and cTnI to cTnC. Nuclear magnetic resonance chemical
shift and relaxation data provided evidence that the cNTnC hydrophobic
core is more exposed with the L48Q variant. Molecular dynamics simulations
suggest that the mutation disrupts a network of crucial hydrophobic
interactions so that the closed form of cNTnC is destabilized. The
findings emphasize the importance of cNTnC’s conformation in
the regulation of contraction and suggest that mutations in cNTnC
that alter myofilament Ca<sup>2+</sup> sensitivity can do so by modulating
Ca<sup>2+</sup> and cTnI binding