Localization of regions of troponin I important in deactivation of cardiac myofilaments by acidic pH.
Ca2+-activation of cardiac muscle myofilaments is more sensitive to depression by acidic pH than is the case with skeletal myofilaments. We tested the hypothesis that this difference is related to specific regions of the TnI (troponin I) isoforms in these muscles. We exchanged native Tn complex in detergent-extracted fiber bundles from mouse ventricles with Tn containing various combinations of fast (fsTnI) or slow skeletal (ssTnI) complexed with either cardiac TnC (cTnC) or fsTnC, and with cTnC complexed with the following chimeras: (1) fsTnI N-terminal region (fN) plus cTnI inhibitory peptide (cIp) and cTnI C-terminal region (cC); and (2) cTnI N-terminal region (cN)-cIp-fsTnI C-terminal region (fC). We determined the change in half maximal Ca2+(DeltaEC50) for tension activation at pH 7.0 and pH 6.5. Similar DeltaEC50 values were obtained for unextracted controls (5.53+/-0.30 microm), for preparations containing cTnI-cTnC (5.74+/-0.40 microm), and preparations exchanged with cTnI-fsTnC (5.63+/-0.40 microm). However, replacement of cTnI with fsTnI significantly decreased DeltaEC50 to 3.95+/-0.17 microm. Replacement of cTnI with ssTnI also significantly depressed DeltaEC50 to 2.07+/-0.15 microm. Results of studies using the chimeras demonstrated that the C-terminal domains of cTnI and fsTnI are responsible for these differences. This conclusion also fits with data from experiments in which we measured Ca2+-binding to the regulatory site of cTnC in binary complexes containing cTnC with cTnI, fsTnI, or the chimeras. Our results localize a region of TnI important in effects of acidosis on cardiac myofilaments and extend our earlier data indicating that C-terminal regions of cTnI outside the Ip are critical for activation by Ca2+.
Li
G
G
Department of Physiology and Biophysics, College of Medicine, Chicago, IL 60612, USA.
Martin
A F
AF
Solaro
R J
RJ
D000136:Acid-Base Equilibrium / Q000502:physiology*; D008841:Actin Cytoskeleton / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D020816:Amino Acid Motifs / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D009206:Myocardium / Q000737:chemistry / Q000378:metabolism*; D009210:Myofibrils / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019209:Troponin C / Q000737:chemistry / Q000378:metabolism*; D019210:Troponin I / Q000737:chemistry / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D019209:Troponin C; D019210:Troponin I; D002118:Calcium