title: Allosteric Transmission along a Loosely Structured Backbone Allows a Cardiac Troponin C Mutant to Function with Only One Ca2+Ion.
abstract: Hypertrophic cardiomyopathy (HCM) is one of the most common cardiomyopathies and a major cause of sudden death in young athletes. The Ca2+sensor of the sarcomere, cardiac troponin C (cTnC), plays an important role in regulating muscle contraction. Although several cardiomyopathy-causing mutations have been identified in cTnC, the limited information about their structural defects has been mapped to the HCM phenotype. Here, we used high-resolution electron-spray ionization mass spectrometry (ESI-MS), Carr-Purcell-Meiboom-Gill relaxation dispersion (CPMG-RD), and affinity measurements of cTnC for the thin filament in reconstituted papillary muscles to provide evidence of an allosteric mechanism in mutant cTnC that may play a role to the HCM phenotype. We showed that the D145E mutation leads to altered dynamics on a μs-ms time scale and deactivates both of the divalent cation-binding sites of the cTnC C-domain. CPMG-RD captured a low populated protein-folding conformation triggered by the Glu-145 replacement of Asp. Paradoxically, although D145E C-domain was unable to bind Ca2+, these changes along its backbone allowed it to attach more firmly to thin filaments than the wild-type isoform, providing evidence for an allosteric response of the Ca2+-binding site II in the N-domain. Our findings explain how the effects of an HCM mutation in the C-domain reflect up into the N-domain to cause an increase of Ca2+affinity in site II, thus opening up new insights into the HCM phenotype.
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil.
authors.forename: Mayra de A
authors.initials: MA
authors.lastname: Marques
authors.affiliation: the Department of Biomedical Sciences, College of Medicine, Florida State University, Tallahassee, Florida 32304.
authors.forename: Jose Renato
authors.initials: JR
authors.lastname: Pinto
authors.affiliation: the Departamento de Química, Instituto de Ciências Exatas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais 31270-901, Brazil, and.
authors.forename: Adolfo H
authors.initials: AH
authors.lastname: Moraes
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil.
authors.forename: Anwar
authors.initials: A
authors.lastname: Iqbal
authors.affiliation: the Departamento de Bioquímica e Imunologia, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais 31270-901, Brazil.
authors.forename: Mariana T Q
authors.initials: MT
authors.lastname: de Magalhães
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil.
authors.forename: Jamila
authors.initials: J
authors.lastname: Monteiro
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil.
authors.forename: Murilo M
authors.initials: MM
authors.lastname: Pedrote
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil.
authors.forename: Martha M
authors.initials: MM
authors.lastname: Sorenson
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil, jerson@bioqmed.ufrj.br.
authors.forename: Jerson L
authors.identifier: 0000-0001-9523-9441
authors.initials: JL
authors.lastname: Silva
authors.affiliation: From the Programa de Biologia Estrutural, Instituto de Bioquímica Médica, Instituto Nacional de Biologia Estrutural e Bioimagem, Centro Nacional de Ressonância Magnética Nuclear Jiri Jonas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902, Brazil, gaugusto@bioqmed.ufrj.br.
authors.forename: Guilherme A P
authors.initials: GA
authors.lastname: de Oliveira
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D002312:Cardiomyopathy, Hypertrophic / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D009154:Mutation; D009206:Myocardium / Q000378:metabolism*; D011487:Protein Conformation; D051381:Rats; D017208:Rats, Wistar; D013057:Spectrum Analysis / Q000379:methods; D019209:Troponin C / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D019209:Troponin C
keywords: calcium-binding protein; cardiomyopathy; nuclear magnetic resonance (NMR); protein structure; small-angle X-ray scattering (SAXS); troponin