The heart-specific NH₂-terminal extension regulates the molecular conformation and function of cardiac troponin I

Abstract

Akhter S, Zhang Z, Jin JP. The heart-specific NH2-terminal extension regulates the molecular conformation and function of cardiac troponin I. Am J Physiol Heart Circ Physiol 302: H923-H933, 2012. First published December 2, 2011; doi:10.1152/ajpheart.00637.2011.-In addition to the core structure conserved in all troponin I isoforms, cardiac troponin I (cTnI) has an ϳ30 amino acids NH 2-terminal extension. This peptide segment is a heart-specific regulatory structure containing two Ser residues that are substrates of PKA. Under ␤-adrenergic regulation, phosphorylation of cTnI in the NH 2-terminal extension increases the rate of myocardial relaxation. The NH 2-terminal extension of cTnI is also removable by restrictive proteolysis to produce functional adaptation to hemodynamic stresses. The molecular mechanism for the NH 2-terminal modifications to regulate the function of cTnI is not fully understood. In the present study, we tested a hypothesis that the NH 2-terminal extension functions by modulating the conformation of other regions of cTnI. Monoclonal antibody epitope analysis and protein binding experiments demonstrated that deletion of the NH 2-terminal segment altered epitopic conformation in the middle, but not COOH-terminal, region of cTnI. PKA phosphorylation produced similar effects. This targeted longrange conformational modulation corresponded to changes in the binding affinities of cTnI for troponin T and for troponin C in a Ca 2ϩ -dependent manner. The data suggest that the NH2-terminal extension of cTnI regulates cardiac muscle function through modulating molecular conformation and function of the core structure of cTnI.troponin I phosphorylation; proteolytic NH 2-terminal truncation; TnITnT interface; cardiac muscle regulation; protein conformation analysis THE CONTRACTION OF CARDIAC muscle is regulated by binding of cytosolic Ca 2ϩ to troponin, which activates cross bridge cycling between sarcomeric myosin and actin filaments. The troponin complex consists of three protein subunits: the Ca 2ϩ -binding subunit troponin C (TnC), the tropomyosin-binding subunit troponin T (TnT), and the inhibitory subunit troponin I (TnI) (19). The function of TnI is essential to cardiac muscle contraction (34).Three homologous genes are present in vertebrate species encoding the fast skeletal muscle, slow skeletal muscle, and cardiac isoforms of TnI (20,29). Cardiac TnI (cTnI) is the newest member evolved in the family of TnI isoform genes (7). In addition to the ϳ180 amino acids core structure that is highly conserved in the three TnI isoforms in all vertebrates, cTnI has a unique NH 2 -terminal extension of ϳ30 amino acids, which is not found in the two skeletal muscle TnI isoforms.Showing functional conservation and exchangeability among the TnI isoforms, embryonic cardiac muscle utilizes solely slow skeletal muscle TnI. During development, it is completely replaced by cTnI in the adult heart (22, 36). Therefore, the NH 2 -terminal extension of cTnI is not an essential structure for the basic contractility of c...

Increased High-Sensitivity Troponin-T Levels Are Associated with Mortality After Ischemic Stroke

“…This specificity is present in early developmental stages in the embryo, where cardiac troponins are only expressed in beating heart tissue (Filipczyk et al 2007) and are mediated by a number of mechanisms, including specific promoters for cardiac troponin (Harlan et al 2008) and an amino acid sequence different from other troponin proteins (Akhter et al 2012). However, later in life, the troponin complex is prevalent in other tissues as well, and there is evidence suggesting that under certain conditions, genes encoding the cardiac troponin complex (type c) can undergo activation and be expressed even in the brain (Lyckman et al 2008).…”

TNNI1, TNNI2 and TNNI3: Evolution, regulation, and protein structure–function relationships

“…While the N-terminal extension of cTnI does not have definitive interaction with other known myofilament proteins, it plays a role in modulating the conformation of other regions of the cTnI molecule (Akhter et al, 2012). …”

“…The N-terminal extension contains the key PKA phosphorylation sites and plays a role in modulating the overall molecular conformation and function of cTnI (Akhter et al, 2012). Different from the C-terminal truncation, a selective removal of the N-terminal extension of cTnI through restrictive proteolysis occurs as a regulatory mechanism in cardiac adaptation in physiological and pathological stress conditions.…”

Effect of N-Terminal Extension of Cardiac Troponin I on the Ca2+ Regulation of ATP Binding and ADP Dissociation of Myosin II in Native Cardiac Myofibrils

“…cTnI also contains a calcium dependent regulatory region and a coiled-coil region (the I–T arm) that interacts with TnT and transmits calcium signals in the troponin complex 12–15 . Recent studies have suggested that the N-terminal extension of cTnI regulates the function of cTnI through long range conformational effect on the I–T interface 15, 16 .…”