EC:3.4.21.69 - FACTA Search

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Query: EC:3.4.21.69 (APC)
16,337 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Myosin-binding-protein C (MyBP-C) is a myosin-associated protein of unknown function found in the cross-bridge-bearing zone (C region) of A bands in striated muscle. Using a cDNA clone encoding the fast-type isoform of chicken MyBP-C, we screened a human fetal muscle cDNA library and isolated clones encoding the full-length human fast-type isoform of MyBP-C. cDNA clones encoding the slow-type isoform of human MyBP-C, were also isolated and fully sequenced. Northern-blot analysis demonstrated skeletal muscle-specific expression of these gene products. Using human/hamster somatic-cell hybrids, we were able to map the slow-type MyBP-C to human chromosome 12, and the fast-type MyBP-C to chromosome 19. The cDNA for human fast-type MyBP-C encodes a polypeptide of 1142 amino acids with an expected molecular mass of 128.1 kDa. Comparison of this cDNA with other members of the MyBP family reveals extensive primary-sequence conservation. Each MyBP-C contains seven immunoglobulin C2 motifs and three fibronectin type-III repeats in the arrangement C2-C2-C2-C2-C2-III-III-C2-III-C2. Regions of high identity shared by the chicken and the two human proteins are not restricted to the immunoglobulin and fibronectin motifs. Sequence comparison of all three proteins has allowed us to map a highly conserved region between the first and second C2 motifs, the only large spacer sequence present between motifs in these proteins.
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PMID:Complete sequence of human fast-type and slow-type muscle myosin-binding-protein C (MyBP-C). Differential expression, conserved domain structure and chromosome assignment. 837

Myosin binding protein C (MyBP-C) is a major myofibril-associated protein in cardiac muscle which is subject to reversible phosphorylation. Cardiac MyBP-C is a substrate in vivo and in vitro for cAMP-dependent protein kinase (PKA) and calcium/phospholipid-dependent protein kinase (PKC). Chicken cardiac MyBP-C was phosphorylated by PKA to 3.0 mol phosphate/mol and by PKC to 2.0 mol phosphate/mol. Tryptic phosphopeptides from MyBP-C were purified by successive iron iminodiacetate column chromatography and reversed-phase high-performance liquid chromatography. Three phosphopeptides purified from PKA-phosphorylated MyBP-C contained phosphoserine [T1, (RTS[P]LAGGGR) and T2, (KRDS[P]FLR)] or phosphothreonine (CT3, MT[P]SAFL). PKC phosphorylated two of the same sites (T1 and T2) as PKA and an additional site [T2a (TGTTYKPPS[P]YK)]. PKA phosphorylation sites corresponding to peptides T1, T2, and T3 were identified in the N-terminus of the cDNA deduced amino acid sequence (S265, S300, and T274, respectively). The PKC-specific site in peptide T2a was at position S1169. cDNA clones encoding rat cardiac MyBP-C were isolated, and the segment corresponding to PKA and major PKC phosphorylation sites was sequenced. Chicken cardiac MyBP-C has a threonine at position 274 (CT3), whereas rat cardiac MyBP-C has a serine at the corresponding position. Only chicken cardiac MyBP-C had a phosphorylatable residue at the position corresponding to S1169. All of the cardiac MyBP-C phosphorylation sites are absent in known sequences of skeletal muscle MyBP-C isoforms.
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PMID:Cardiac myosin-binding protein C (MyBP-C): identification of protein kinase A and protein kinase C phosphorylation sites. 978 45

Myosin binding protein C (MyBP-C) is one of a group of myosin binding proteins that are present in the myofibrils of all striated muscle. The protein is found at 43-nm repeats along 7 to 9 transverse lines in a portion of the A band where crossbridges are found (C zone). MyBP-C contains myosin and titin binding sites at the C terminus of the molecule in all 3 of the isoforms (slow skeletal, fast skeletal, and cardiac). The cardiac isoform also includes a series of residues that contain 3 phosphorylatable sites and an additional immunoglobulin module at the N terminus that are not present in skeletal isoforms. The following 2 major functions of MyBP-C have been suggested: (1) a role in the formation of the sarcomeric myofibril as a result of binding to myosin and titin and (2) in the case of the cardiac isoform, regulation of contraction through phosphorylation. The first is supported by the demonstrated effect of MyBP-C on the packing of myosin in the thick filament, the coincidence of appearance of sarcomeres and MyBP-C during myofibrillogenesis, and the defective formation of sarcomeres when the titin and/or myosin binding sites of MyBP-C are missing. The second is supported by the specific phosphorylation sites in cardiac MyBP-C, the presence in the thick filament of an enzyme specific for MyBP-C phosphorylation, the alteration of thick filament structure by MyBP-C phosphorylation, and the accompaniment of MyBP-C phosphorylation with all major physiological mechanisms of modulation of inotropy in the heart.
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PMID:Cardiac myosin binding protein C. 1034 86

Myosin binding protein C is a protein of the myosin filaments of striated muscle which is expressed in isoforms specific for cardiac and skeletal muscle. The cardiac isoform is phosphorylated rapidly upon adrenergic stimulation of myocardium by cAMP-dependent protein kinase, and together with the phosphorylation of troponin-I and phospholamban contributes to the positive inotropy that results from adrenergic stimulation of the heart. Cardiac myosin binding protein C is phosphorylated by cAMP-dependent protein kinase on three sites in a myosin binding protein C specific N-terminal domain which binds to myosin-S2. This interaction with myosin close to the motor domain is likely to mediate the regulatory function of the protein. Cardiac myosin binding protein C is a common target gene of familial hypertrophic cardiomyopathy and most mutations encode N-terminal subfragments of myosin binding protein C. The understanding of the signalling interactions of the N-terminal region is therefore important for understanding the pathophysiology of myosin binding protein C associated cardiomyopathy. We demonstrate here by cosedimentation assays and isothermal titration calorimetry that the myosin-S2 binding properties of the myosin binding protein C motif are abolished by cAMP-dependent protein kinase-mediated tris-phosphorylation, decreasing the S2 affinity from a Kd of approximately 5 microM to undetectable levels. We show that the slow and fast skeletal muscle isoforms are no cAMP-dependent protein kinase substrates and that the S2 interaction of these myosin binding protein C isoforms is therefore constitutively on. The regulation of cardiac contractility by myosin binding protein C therefore appears to be a 'brake-off' mechanism that will free a specific subset of myosin heads from sterical constraints imposed by the binding to the myosin binding protein C motif.
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PMID:cAPK-phosphorylation controls the interaction of the regulatory domain of cardiac myosin binding protein C with myosin-S2 in an on-off fashion. 1040 55

Myosin binding protein C (MyBP-C) is an integral part of the striated muscle sarcomere. As is the case for other sarcomeric genes in human populations, multiple mutations within the gene have been linked to familial hypertrophic cardiomyopathy. Although some MyBP-C lesions are the result of missense mutations, most show truncated polypeptides lacking either the myosin or myosin and titin binding sites. Previously, we generated transgenic (TG) mice with cardiac-specific expression of a MyBP-C mutant lacking the myosin and titin binding domains. Surprisingly, the mutant protein was stable and made up a majority of the MyBP-C species, with concomitant reductions in endogenous MyBP-C such that overall MyBP-C stoichiometry was conserved. In the present study, we created a second series of TG mice that express, in the heart, a mutant MyBP-C lacking only the myosin binding site. In contrast to the previous data for the MyBP-C lacking both titin and myosin binding sites, only very modest levels of protein were found, consistent with data obtained from human biopsies in which mutated MyBP-C could not be detected. Despite normal levels of wild-type MyBP-C, there were significant changes in the structure and ultrastructure of the heart. Fiber mechanics showed decreased unloading shortening velocity, maximum shortening velocity, and relative maximal power output.
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PMID:In vivo modeling of myosin binding protein C familial hypertrophic cardiomyopathy. 1053 52

Myosin binding protein C (MyBP-C) is one of the major sarcomeric proteins involved in the pathophysiology of familial hypertrophic cardiomyopathy (FHC). The cardiac isoform is tris-phosphorylated by cAMP-dependent protein kinase (cAPK) on beta-adrenergic stimulation at a conserved N-terminal domain (MyBP-C motif), suggesting a role in regulating positive inotropy mediated by cAPK. Recent data show that the MyBP-C motif binds to a conserved segment of sarcomeric myosin S2 in a phosphorylation-regulated way. Given that most MyBP-C mutations that cause FHC are predicted to result in N-terminal fragments of the protein, we investigated the specific effects of the MyBP-C motif on contractility and its modulation by cAPK phosphorylation. The diffusion of proteins into skinned fibers allows the investigation of effects of defined molecular regions of MyBP-C, because the endogenous MyBP-C is associated with few myosin heads. Furthermore, the effect of phosphorylation of cardiac MyBP-C can be studied in a defined unphosphorylated background in skeletal muscle fibers only. Triton skinned fibers were tested for maximal isometric force, Ca(2+)/force relation, rigor force, and stiffness in the absence and presence of the recombinant cardiac MyBP-C motif. The presence of unphosphorylated MyBP-C motif resulted in a significant (1) depression of Ca(2+)-activated maximal force with no effect on dynamic stiffness, (2) increase of the Ca(2+) sensitivity of active force (leftward shift of the Ca(2+)/force relation), (3) increase of maximal rigor force, and (4) an acceleration of rigor force and rigor stiffness development. Tris-phosphorylation of the MyBP-C motif by cAPK abolished these effects. This is the first demonstration that the S2 binding domain of MyBP-C is a modulator of contractility. The anchorage of the MyBP-C motif to the myosin filament is not needed for the observed effects, arguing that the mechanism of MyBP-C regulation is at least partly independent of a "tether," in agreement with a modulation of the head-tail mobility. Soluble fragments occurring in FHC, lacking the spatial specificity, might therefore lead to altered contraction regulation without affecting sarcomere structure directly.
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PMID:Myosin binding protein C, a phosphorylation-dependent force regulator in muscle that controls the attachment of myosin heads by its interaction with myosin S2. 1062 98

Myosin-binding protein C (MyBP-C) is thought to play structural and/or regulatory role in striated muscles. The cardiac isoform of MyBP-C is one of the disease genes associated with familial hypertrophic cardiomyopathy and most of the mutations produce COOH truncated proteins. In order to determine the consequences of these mutations on myosin filament organization, we have characterized the effect of a 52-kDa NH2-terminal peptide of human cardiac MyBP-C on the alpha-myosin heavy chain (alpha-MyHC) filament organization. This peptide lacks the COOH-terminal MyHC-binding site and retains the two MyHC-binding domains located in the N-terminal part of MyBP-C. For this characterization, cDNA constructs (rat alpha-MyHC, full-length and truncated human cardiac MyBP-C) were transiently expressed singly or in pairwise combination in COS cells. In conformity with previous works performed on the skeletal isoform of MyBP-C, we observed that full-length cardiac MyBP-C organizes the MyHC into dense structures of uniform width. While the truncated protein is stable and can interact with MyHC in COS cells, it does not result in the same organization of sarcomeric MyHC that is seen with the full-length MyBP-C. These results suggest that the presence of truncated cardiac MyBP-C could, at least partly, disorganize the sarcomeric structure in patients with familial hypertrophic cardiomyopathy.
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PMID:COOH-terminal truncated human cardiac MyBP-C alters myosin filament organization. 1129 12

Four and a half LIM protein 1 (FHL1/SLIM1) is highly expressed in skeletal and cardiac muscle; however, the function of FHL1 remains unknown. Yeast two-hybrid screening identified slow type skeletal myosin-binding protein C as an FHL1 binding partner. Myosin-binding protein C is the major myosin-associated protein in striated muscle that enhances the lateral association and stabilization of myosin thick filaments and regulates actomyosin interactions. The interaction between FHL1 and myosin-binding protein C was confirmed using co-immunoprecipitation of recombinant and endogenous proteins. Recombinant FHL2 and FHL3 also bound myosin-binding protein C. FHL1 impaired co-sedimentation of myosin-binding protein C with reconstituted myosin filaments, suggesting FHL1 may compete with myosin for binding to myosin-binding protein C. In intact skeletal muscle and isolated myofibrils, FHL1 localized to the I-band, M-line, and sarcolemma, co-localizing with myosin-binding protein C at the sarcolemma in intact skeletal muscle. Furthermore, in isolated myofibrils FHL1 staining at the M-line appeared to extend partially into the C-zone of the A-band, where it co-localized with myosin-binding protein C. Overexpression of FHL1 in differentiating C2C12 cells induced "sac-like" myotube formation (myosac), associated with impaired Z-line and myosin thick filament assembly. This phenotype was rescued by co-expression of myosin-binding protein C. FHL1 knockdown using RNAi resulted in impaired myosin thick filament formation associated with reduced incorporation of myosin-binding protein C into the sarcomere. This study identified FHL1 as a novel regulator of myosin-binding protein C activity and indicates a role for FHL1 in sarcomere assembly.
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PMID:Four and a half LIM protein 1 binds myosin-binding protein C and regulates myosin filament formation and sarcomere assembly. 1640 97

Myosin heads interacting with actin filaments, a process fueled by MgATP and regulated by calcium, powers the pump-like action of the human heart. Hydrolysis of MgATP, the competition between MgATP, its products of hydrolysis, and actin for binding to myosin, and the sequence of shifting affinities in that competition, constitute the central mechanism of muscular contraction. The force, work, and power produced during the cardiac cycle stems from an isomerization of the myosin head that is closely associated with strong binding of myosin to actin and release of phosphate. While fluctuations of intracellular [Ca2+] bound to troponin and related shifts in tropomyosin on the thin filaments regulate the number of crossbridges on a beat-to-beat basis, the oscillatory work produced is augmented by a delayed force response to stretch that develops during diastole. This stretch-activated myogenic response is facilitated by specialized myofilament structures, including actin-binding portions of the myosin essential light chain and myosin binding protein C, which are thought to guide and orient the myosin head or enhance thin filament activation. Phosphorylation of the myosin regulatory light chain, myosin binding protein C, and troponin T also assist in this regard. Animal models show isoform shifts in myosin and other myofibrillar proteins have major effects on power output, but isoform shifts in human myocardium are modest at best and are therefore likely to play only a minor role in modulating crossbridge kinetics compared to disease-related post-translational modifications of the contractile proteins and to changes in their chemical environment.
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PMID:Kinetics and energetics of the crossbridge cycle. 1641 41

Myosin II is the major component of the muscle thick filament. It consists of two N-terminal S1 subfragments ("heads") connected to a long dimeric coiled-coil rod. The rod is in itself twofold symmetric, but in the filament, the two heads point away from the filament surface and are therefore not equivalent. This breaking of symmetry requires the initial section of the rod, subfragment 2 (S2), to be relatively flexible. S2 is an important functional element, involved in various mechanisms by which the activity of smooth and striated muscle is regulated. We have determined crystal structures of the 126 N-terminal residues of S2 from human cardiac beta-myosin II (S2-Delta), of both WT and the disease-associated E924K mutant. S2-Delta is a straight parallel dimeric coiled coil, but the N terminus of one chain is disordered in WT-S2-Delta due to crystal contacts, indicative of unstable local structure. Bulky noncanonical side chains pack into a/d positions of S2-Delta's N terminus, leading to defined local asymmetry and axial stagger, which could induce nonequivalence of the S1 subfragments. Additionally, S2 possesses a conserved charge distribution with three prominent rings of negative potential within S2-Delta, the first of which may provide a binding interface for the "blocked head" of smooth muscle myosin in the OFF state. The observation that many disease-associated mutations affect the second negatively charged ring further suggests that charge interactions play an important role in regulation of cardiac muscle activity through myosin-binding protein C.
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PMID:Crystal structures of human cardiac beta-myosin II S2-Delta provide insight into the functional role of the S2 subfragment. 1709 4

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