Modification of the kinetic parameters of aldolase on binding to the actin-containing filaments of skeletal muscle.
Ontology highlight
ABSTRACT:
SUBMITTER: Walsh TP
PROVIDER: S-EPMC1164882 | biostudies-other | 1977 Jul
REPOSITORIES: biostudies-other
Similar Datasets
Project description:The interactions of aldolase with regulatory proteins of rabbit skeletal muscle were investigated by moving-boundary electrophoresis. A salt-dependent interaction of troponin, tropomyosin and the tropomyosin-troponin complex with aldolase was detected, the tropomyosin-troponin complex displaying a greater affinity for the enzyme than did either regulatory protein alone. The results indicate that aldolase possesses multiple binding sites (three or more) for these muscle proteins. Quantitative studies of the binding of aldolase to actin-containing filaments showed the interaction to be influenced markedly by the presence of these muscle regulatory proteins on the filaments. In imidazole/HCl buffer, I 0.088, pH 6.8, aldolase binds to F-actin with an affinity constant of 2 x 10(5) M-1 and a stoicheiometry of one tetrameric aldolase molecule per 14 monomeric actin units. Use of F-actin-tropomyosin as adsorbent results in a doubling of the stoicheiometry without significant change in the intrinsic association constant. With F-actin-tropomyosin-troponin a lower binding constant (6 x 10(4) M-1) but even greater stoicheiometry (4:14 actin units) are observed. The presence of Ca2+ (0.1 mM) decreases this stoicheiometry to 3:14 without affecting significantly the magnitude of the intrinsic binding constant.
| S-EPMC1161506 | biostudies-other
Project description:Electron-microscopy observation show that when aldolase binds to F-actin or F-actin-tropomyosin, highly ordered paracrystalline structures are formed consisting of tightly packed filament bundles cross-banded at 36 nm intervals. Morphologically different paracrystalline arrays are formed between aldolase and F-actin-tropomyosin-troponin. The filament bundles are far more extensive and are characterized by a prominent cross-striation at 38nm intervals. It is suggested that this reflects an interaction between troponin and aldolase.
| S-EPMC1164183 | biostudies-other
Project description:Previously reported results of equilibrium-partition experiments on the interaction of aldolase with actin-containing filaments [Walsh, Winzor, Clarke, Masters & Morton (1980) Biochem. J. 186, 89-98] have been subjected to a more rigorous theoretical analysis involving consideration of the consequences of cross-linking interactions between enzyme and filament. The experimental results obtained with F-actin-tropomyosin are best described by a model with one binding site per heptameric repeat unit of filament and a value of 39000 M-1 for the site binding constant, k. Similar analyses of the influence of Ca2+ on aldolase binding to F-actin--tropomyosin--troponin substantiate the existence of two equivalent binding sites (k = 14900 M-1) for the enzyme on each repeat unit of the thin filament. The Ca2+-sensitivity of this interaction reflects either a decrease in the strength of aldolase binding to these two sites (k = 8200 M-1) or the elimination of one site.
| S-EPMC1162885 | biostudies-other
Project description:Electron micrographs of the paracrystals formed when fructose bisphosphate aldolase (EC 4.1.2.13) is added to actin-containing filaments were analysed by computer methods so that ultrastructural changes could be correlated with the various stoicheiometries of binding determined in the preceding paper [Walsh, Winzor, Clarke, Masters & Morton (1980) Biochem. J. 186, 89-98]. Paracrystals formed with aldolase and either F-actin or F-actin-tropomyosin have a single light transverse band every 38 nm, which is due to aldolase molecules cross-linking the filaments. In contrast, the paracrystals formed between aldolase and F-actin-tropomyosin-troponin filaments show two transverse bands every 38 nm: a major band, interpreted as aldolase binding to troponin, and a minor band, interpreted as aldolase cross-linking the filaments. The intensity of the minor band varies with Ca2+ concentration, being greatest when the Ca2+ concentration is low. A model for the different paracrystal structures which relates the various patterns and binding stoicheiometries to structural changes in the actin-containing filaments is proposed.
| S-EPMC1161507 | biostudies-other
Project description:The core of skeletal muscle Z-discs consists of actin filaments from adjacent sarcomeres that are cross-linked by ?-actinin homodimers. Z-disc-associated, alternatively spliced, PDZ motif-containing protein (ZASP)/Cypher interacts with ?-actinin, myotilin, and other Z-disc proteins via the PDZ domain. However, these interactions are not sufficient to maintain the Z-disc structure. We show that ZASP directly interacts with skeletal actin filaments. The actin-binding domain is between the modular PDZ and LIM domains. This ZASP region is alternatively spliced so that each isoform has unique actin-binding domains. All ZASP isoforms contain the exon 6-encoded ZASP-like motif that is mutated in zaspopathy, a myofibrillar myopathy (MFM), whereas the exon 8-11 junction-encoded peptide is exclusive to the postnatal long ZASP isoform (ZASP-L?ex10). MFM is characterized by disruption of skeletal muscle Z-discs and accumulation of myofibrillar degradation products. Wild-type and mutant ZASP interact with ?-actin, ?-actinin, and myotilin. Expression of mutant, but not wild-type, ZASP leads to Z-disc disruption and F-actin accumulation in mouse skeletal muscle, as in MFM. Mutations in the actin-binding domain of ZASP-L?ex10, but not other isoforms, cause disruption of the actin cytoskeleton in muscle cells. These isoform-specific mutation effects highlight the essential role of the ZASP-L?ex10 isoform in F-actin organization. Our results show that MFM-associated ZASP mutations in the actin-binding domain have deleterious effects on the core structure of the Z-discs in skeletal muscle.
| S-EPMC4036366 | biostudies-literature
Project description:Clathrin plaques are stable features of the plasma membrane observed in several cell types. They are abundant in muscle, where they localize at costameres that link the contractile apparatus to the sarcolemma and connect the sarcolemma to the basal lamina. Here, we show that clathrin plaques and surrounding branched actin filaments form microdomains that anchor a three-dimensional desmin intermediate filament (IF) web. Depletion of clathrin plaque and branched actin components causes accumulation of desmin tangles in the cytoplasm. We show that dynamin 2, whose mutations cause centronuclear myopathy (CNM), regulates both clathrin plaques and surrounding branched actin filaments, while CNM-causing mutations lead to desmin disorganization in a CNM mouse model and patient biopsies. Our results suggest a novel paradigm in cell biology, wherein clathrin plaques act as platforms capable of recruiting branched cortical actin, which in turn anchors IFs, both essential for striated muscle formation and function.
| S-EPMC6589689 | biostudies-literature
Project description:Several actin-binding proteins (ABPs) phase separate to form condensates capable of curating the actin network shapes. Here, we use computational modeling to understand the principles of actin network organization within VASP condensate droplets. Our simulations reveal that the different actin shapes, namely shells, rings, and mixture states are highly dependent on the kinetics of VASP-actin interactions, suggesting that they arise from kinetic trapping. Specifically, we show that reducing the residence time of VASP on actin filaments reduces degree of bundling, thereby promoting assembly of shells rather than rings. We validate the model predictions experimentally using a VASP-mutant with decreased bundling capability. Finally, we investigate the ring opening within deformed droplets and found that the sphere-to-ellipsoid transition is favored under a wide range of filament lengths while the ellipsoid-to-rod transition is only permitted when filaments have a specific range of lengths. Our findings highlight key mechanisms of actin organization within phase-separated ABPs.
| S-EPMC11009352 | biostudies-literature
Project description:Cytokeratins 8 and 19 concentrate at costameres of striated muscle and copurify with the dystrophin-glycoprotein complex, perhaps through the interaction of the cytokeratins with the actin-binding domain of dystrophin. We overexpressed dystrophin's actin-binding domain (Dys-ABD), K8 and K19, as well as closely related proteins, in COS-7 cells to assess the basis and specificity of their interaction. Dys-ABD alone associated with actin microfilaments. Expressed with K8 and K19, which form filaments, Dys-ABD associated preferentially with the cytokeratins. This interaction was specific, as the homologous ABD of betaI-spectrin failed to interact with K8/K19 filaments, and Dys-ABD did not associate with desmin or K8/K18 filaments. Studies in COS-7 cells and in vitro showed that Dys-ABD binds directly and specifically to K19. Expressed in muscle fibers in vivo, K19 accumulated in the myoplasm in structures that contained dystrophin and spectrin and disrupted the organization of the sarcolemma. K8 incorporated into sarcomeres, with no effect on the sarcolemma. Our results show that dystrophin interacts through its ABD with K19 specifically and are consistent with the idea that cytokeratins associate with dystrophin at the sarcolemma of striated muscle.
| S-EPMC1196337 | biostudies-literature
Project description:Skeletal muscle actin mice (Crawford et al., (2002) Mol Cell Biol 22, 5587) were crossed with cardiac actin transgenic mice (termed "ACTC^Coco" or "Coco" for short), to produce mice that had cardiac actin instead of skeletal muscle actin in their skeletal muscles (termed "ACTC^Co/KO" or for short "Coco/KO"). Microarray analysis using the Illumina mouse-6 v1.1 expression beadchip was performed on RNA extraced from the soleus muscle of Coco/KO mice and wildtype mice, to confirm the swith in actin isoform expression, and to determine what other differences might exist between wildtype mice and the Coco/KO mice. Keywords: genetic modification 3 RNA samples (each being the pool of two individual samples extracted from different soleus muscles from different individual mice) per genotype (either wildtype or Coco/KO) were used. The total 6 RNA samples were processed using an Illumina mouse-6 v1.1expression beadchip and then the differentially expressed genes determined.
2009-05-24 | E-GEOD-12882 | biostudies-arrayexpress
Project description:Skeletal muscle actin mice (Crawford et al., (2002) Mol Cell Biol 22, 5587) were crossed with cardiac actin transgenic mice (termed "ACTC^Coco" or "Coco" for short), to produce mice that had cardiac actin instead of skeletal muscle actin in their skeletal muscles (termed "ACTC^Co/KO" or for short "Coco/KO"). Microarray analysis using the Illumina mouse-6 v1.1 expression beadchip was performed on RNA extraced from the soleus muscle of Coco/KO mice and wildtype mice, to confirm the swith in actin isoform expression, and to determine what other differences might exist between wildtype mice and the Coco/KO mice. Keywords: genetic modification
2009-05-25 | GSE12882 | GEO