Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.4.1 (myosin ATPase)
1,140 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Calcium binding proteins mediate a large number of cellular processes. These processes respond to micromolar fluctuations of cytosolic calcium in the presence of a large excess of magnesium. The metal binding sites present in these proteins are either calcium-specific (regulatory sites) or capable of binding both calcium and magnesium (structural sites). Using site-directed mutagenesis we were able to convert the single Ca2+/Mg2+ site present in chicken smooth muscle myosin regulatory light chain (RLC) into a Ca(2+)-specific site. The replacement of the aspartic acid present in the 12th position (-Z coordinating position) of the metal binding loop with a glutamic acid increases calcium affinity and abolishes magnesium binding, rendering the site calcium-specific. To explain this observation, we hypothesize that restrictions on the ability of side chains to change conformation, contributing one (for Mg2+ binding) or two (for Ca2+ binding) coordinations could alter the metal specificity in EF-hands. Other mutations which decrease or abolish calcium binding have also been characterized. When used to substitute the endogenous scallop myosin RLC, these mutants were capable of restoring the Ca2+ regulation to the actin-activated myosin ATPase demonstrating that in these hybrid myosins, the regulatory function of the Ca(2+)-specific site (present on the essential light chain) does not depend on the occupancy of the Ca2+/Mg2+ site (present on the regulatory light chain).
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PMID:Determinants of ion specificity on EF-hands sites. Conversion of the Ca2+/Mg2+ site of smooth muscle myosin regulatory light chain into a Ca(2+)-specific site. 789 23

FHC (familial hypertrophic cardiomyopathy) is a heritable form of cardiac hypertrophy caused by mutations in genes encoding sarcomeric proteins. The present study focuses on the A13T mutation in the human ventricular myosin RLC (regulatory light chain) that is associated with a rare FHC variant defined by mid-ventricular obstruction and septal hypertrophy. We generated heart-specific Tg (transgenic) mice with ~10% of human A13T-RLC mutant replacing the endogenous mouse cardiac RLC. Histopathological examinations of longitudinal heart sections from Tg-A13T mice showed enlarged interventricular septa and profound fibrotic lesions compared with Tg-WT (wild-type), expressing the human ventricular RLC, or non-Tg mice. Functional studies revealed an abnormal A13T mutation-induced increase in isometric force production, no change in the force-pCa relationship and a decreased Vmax of the acto-myosin ATPase. In addition, a fluorescence-based assay showed a 3-fold lower binding affinity of the recombinant A13T mutant for the RLC-depleted porcine myosin compared with WT-RLC. These results suggest that the A13T mutation triggers a hypertrophic response through changes in cardiac sarcomere organization and myosin cross-bridge function leading to abnormal remodelling of the heart. The significant functional changes observed, despite a low level of A13T mutant incorporation into myofilaments, suggest a 'poison-peptide' mechanism of disease.
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PMID:Myosin regulatory light chain mutation found in hypertrophic cardiomyopathy patients increases isometric force production in transgenic mice. 2209 67

Phosphorylation of myosin regulatory light chain (RLC) triggers contraction in smooth muscle myocytes. Dephosphorylation of phosphorylated RLC (pRLC) is mediated by myosin RLC phosphatase (MLCP), which is negatively regulated by rho-associated kinase (ROK). We have compared basal and stimulated concentrations of pRLC in myocytes from human coronary artery (hVM), which has a tonic contractile pattern to myocytes from human uterus (hUM), which has a phasic contractile pattern. Our studies reveal fundamental differences between hVM and hUM regarding the mechanisms regulating phosphorylation RLC. Whereas hVM responded to stimulation by phosphorylation of RLC at S19, hUM responded by forming diphosphorylated RLC (at T18 and S19; ppRLC), which, compared to pRLC, causes two to threefold greater activation of myosin ATPase that provides energy to power the contraction. Importantly, the conversion of pRLC to ppRLC is mediated by ROK. In hUM, MLCP has high activity for ppRLC and this is inhibited by ROK through phosphorylation of the substrate targeting subunit (MYPT1) at T853. Inhibitors of ROK significantly reduce contractility in both hVM and hUM. We demonstrated that inhibition of ppRLC in phasic myocytes (hUM) is 100-fold more sensitive to ROK inhibitors than is pRLC in tonic myocytes (hVM). We speculate that these differences in phosphorylation of RLC might reflect evolution of different contractile patterns to perform distinct physiological functions. Furthermore, our data suggest that low concentrations of ROK inhibitors might inhibit uterine contractions with minimal effects on vascular tone, thus posing a novel strategy for prevention or treatment of conditions such as preterm birth.
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PMID:Rho-kinase mediates diphosphorylation of myosin regulatory light chain in cultured uterine, but not vascular smooth muscle cells. 2294 48

Myosin light chain 2 ( MYL2) gene encodes the myosin regulatory light chain (RLC) simultaneously in heart ventricles and in slow-twitch skeletal muscle. Using transgenic mice with cardiac-specific expression of the human R58Q-RLC mutant, we sought to determine whether the hypertrophic cardiomyopathy phenotype observed in papillary muscles (PMs) of R58Q mice is also manifested in slow-twitch soleus (SOL) muscles. Skinned SOL muscles and ventricular PMs of R58Q animals exhibited lower contractile force that was not observed in the fast-twitch extensor digitorum longus muscles of R58Q vs. wild-type-RLC mice, but mutant animals did not display gross muscle weakness in vivo. Consistent with SOL muscle abnormalities in R58Q vs. wild-type mice, myosin ATPase staining revealed a decreased proportion of fiber type I/type II only in SOL muscles but not in the extensor digitorum longus muscles. The similarities between SOL muscles and PMs of R58Q mice were further supported by quantitative proteomics. Differential regulation of proteins involved in energy metabolism, cell-cell interactions, and protein-protein signaling was concurrently observed in the hearts and SOL muscles of R58Q mice. In summary, even though R58Q expression was restricted to the heart of mice, functional similarities were clearly observed between the hearts and slow-twitch skeletal muscle, suggesting that MYL2 mutated models of hypertrophic cardiomyopathy may be useful research tools to study the molecular, structural, and energetic mechanisms of cardioskeletal myopathy associated with myosin RLC.-Kazmierczak, K., Liang, J., Yuan, C.-C., Yadav, S., Sitbon, Y. H., Walz, K., Ma, W., Irving, T. C., Cheah, J. X., Gomes, A. V., Szczesna-Cordary, D. Slow-twitch skeletal muscle defects accompany cardiac dysfunction in transgenic mice with a mutation in the myosin regulatory light chain.
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PMID:Slow-twitch skeletal muscle defects accompany cardiac dysfunction in transgenic mice with a mutation in the myosin regulatory light chain. 3036 66

INS^C94Y transgenic pigs develop a stable diabetic phenotype early after birth and therefore allow studying the influence of hyperglycemia on primary immune cells in an early stage of diabetes mellitus in vivo. Since immune response is altered in diabetes mellitus, with deviant neutrophil function discussed as one of the possible causes in humans and mouse models, we investigated these immune cells in INS^C94Y transgenic pigs and wild type controls at protein level. A total of 2371 proteins were quantified by label-free LC-MS/MS. Subsequent differential proteome analysis of transgenic animals and controls revealed clear differences in protein abundances, indicating a deviant behavior of granulocytes in the diabetic state. Interestingly, abundance of myosin regulatory light chain 9 (MLC-2C) was increased 5-fold in cells of diabetic pigs. MLC-2C directly affects cell contractility by regulating myosin ATPase activity, can act as transcription factor and was also associated with inflammation. It might contribute to impaired neutrophil cell adhesion, migration and phagocytosis. Our study provides novel insights into proteome changes in neutrophils from a large animal model for permanent neonatal diabetes mellitus and points to dysregulation of neutrophil function even in an early stage of this disease. Data are available via ProteomeXchange with identifier PXD017274. SIGNIFICANCE: Our studies provide novel basic information about the neutrophil proteome of pigs and contribute to a better understanding of molecular mechanisms involved in altered immune cell function in an early stage diabetes. We demonstrate proteins that are dysregulated in neutrophils from a transgenic diabetic pig and have not been described in this context so far. The data presented here are highly relevant for veterinary medicine and have translational quality for diabetes in humans.
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PMID:Proteome profile of neutrophils from a transgenic diabetic pig model shows distinct changes. 3247 May 42

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