Phosphorylation-dependent Conformational Transition of the Cardiac Specific N-Extension of Troponin I in Cardiac Troponin
ABSTRACT We present here the solution structure for the bisphosphorylated form of the cardiac N-extension of troponin I (cTnI(1-32)), a region for which there are no previous high-resolution data. Using this structure, the X-ray crystal structure of the cardiac troponin core, and uniform density models of the troponin components derived from neutron contrast variation data, we built atomic models for troponin that show the conformational transition in cardiac troponin induced by bisphosphorylation. In the absence of phosphorylation, our NMR data and sequence analyses indicate a less structured cardiac N-extension with a propensity for a helical region surrounding the phosphorylation motif, followed by a helical C-terminal region (residues 25-30). In this conformation, TnI(1-32) interacts with the N-lobe of cardiac troponin C (cTnC) and thus is positioned to modulate myofilament Ca2+-sensitivity. Bisphosphorylation at Ser23/24 extends the C-terminal helix (residues 21-30) which results in weakening interactions with the N-lobe of cTnC and a re-positioning of the acidic amino terminus of cTnI(1-32) for favorable interactions with basic regions, likely the inhibitory region of cTnI. An extended poly(L-proline)II helix between residues 11 and 19 serves as the rigid linker that aids in re-positioning the amino terminus of cTnI(1-32) upon bisphosphorylation at Ser23/24. We propose that it is these electrostatic interactions between the acidic amino terminus of cTnI(1-32) and the basic inhibitory region of troponin I that induces a bending of cTnI at the end that interacts with cTnC. This model provides a molecular mechanism for the observed changes in cross-bridge kinetics upon TnI phosphorylation.
- SourceAvailable from: Genaro A Ramirez-Correa[Show abstract] [Hide abstract]
ABSTRACT: Global cardiac myofilament proteins phosphorylation levels, and their site-specific stoichiometry, are physiologically and clinically relevant for heart function. Unlike myofilament phosphorylation, other post-translational modifications (PTMs) such as O-GlcNAcylation, are just beginning to gain attention due to their potential physiological and clinical implications. This review will focus on what is currently known about cardiac Troponin I (cTnI) phosphorylation, and on the potential physiological and clinical impact of targeted proteomics including new findings on cTnI sites and stoichiometry. We will then discuss the increasing recognition of other myofilament PTMs functional relevance and the potential of targeted mass spectrometry approaches, particularly multiple reaction monitoring (MRM), for accelerating their systematic characterization. In addition, we will broadly discuss the development and application of MRM to quantitatively assess site-specific PTMs. Finally, we will give an overview of expert's consensus on MRM methods design/validation and best practices to develop MRM assays intended to reach clinical application. The unique ability of MRM and similar methods to identify and quantify cardiac myofilament PTMs is likely to become central in answering important biological questions in the field of cardiac integrative physiology.This article is protected by copyright. All rights reservedPROTEOMICS - CLINICAL APPLICATIONS 08/2014; 8(7-8). DOI:10.1002/prca.201400034 · 2.68 Impact Factor
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ABSTRACT: This review focuses on recent developments in the molecular mechanisms by which Ca activates cardiac sarcomeres and how these mechanisms play out in the cardiac cycle. I emphasize the role of mechanisms intrinsic to the sarcomeres as significant determinants of systolic elastance and ventricular stiffening during ejection. Data are presented supporting the idea that processes intrinsic to the thin filaments may promote cooperative activation of the sarcomeres and be an important factor in maintaining and modifying systolic elastance. Application of these ideas to translational medicine and rationale drug design forms an important rationale for detailed understanding of these processes.BioMed Research International 05/2010; 2010:105648. DOI:10.1155/2010/105648 · 2.71 Impact Factor
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ABSTRACT: Over the 40 years since its discovery, many studies have focused on understanding the role of troponin as a myofilament based molecular switch in regulating the Ca(2+)-dependent activation of striated muscle contraction. Recently, studies have explored the role of cardiac troponin as a target for cardiotonic agents. These drugs are clinically useful for treating heart failure, a condition in which the heart is no longer able to pump enough blood to other organs. These agents act via a mechanism that modulates the Ca(2+)-sensitivity of troponin; such a mode of action is therapeutically desirable because intracellular Ca(2+) concentration is not perturbed, preserving the regulation of other Ca(2+)-based signaling pathways. This review describes molecular details of the interaction of cardiac troponin with a variety of cardiotonic drugs. We present recent structural work that has identified the docking sites of several cardiotonic drugs in the troponin C-troponin I interface and discuss their relevance in the design of troponin based drugs for the treatment of heart disease.Biochemical and Biophysical Research Communications 05/2008; 369(1):88-99. DOI:10.1016/j.bbrc.2007.12.108 · 2.28 Impact Factor