scholarly journals Structural comparison of several actin-binding macromolecules.

1980 ◽  
Vol 85 (2) ◽  
pp. 489-495 ◽  
Author(s):  
J M Tyler ◽  
J M Anderson ◽  
D Branton
Keyword(s):  
Electron Microscopy ◽  
Binding Protein ◽  
Actin Binding ◽  
Erythrocyte Membranes ◽  
Structural Comparison ◽  
Flexible Molecules ◽  
Actin Binding Protein ◽  
Rotary Shadowing

The cytoskeletal components, macrophage actin-binding protein and filamin, were dried from glycerol and examined by low-angle rotary shadowing electron microscopy. Both are elongate, flexible molecules whose general morphologi is similar to that of erythrocyte spectrin. Neither actin-binding protein nor filamin binds to spectrin-depleted erythrocyte membranes.

scholarly journals The architecture of actin filaments and the ultrastructural location of actin-binding protein in the periphery of lung macrophages.

1986 ◽  
Vol 103 (3) ◽  
pp. 1007-1020 ◽  
Author(s):  
J H Hartwig ◽  
P Shevlin
Keyword(s):  
Electron Microscopy ◽  
Critical Point ◽  
Binding Protein ◽  
The Body ◽  
Actin Binding ◽  
Actin Network ◽  
Actin Binding Protein ◽  
Freeze Dried ◽  
Critical Point Drying ◽  
Actin Fiber

A highly branched filament network is the principal structure in the periphery of detergent-extracted cytoskeletons of macrophages that have been spread on a surface and either freeze or critical point dried, and then rotary shadowed with platinum-carbon. This array of filaments completely fills lamellae extended from the cell and bifurcates to form 0.2-0.5 micron thick layers on the top and bottom of the cell body. Reaction of the macrophage cytoskeletons with anti-actin IgG and with anti-IgG bound to colloidal gold produces dense staining of these filaments, and incubation with myosin subfragment 1 uniformly decorates these filaments, identifying them as actin. 45% of the total cellular actin and approximately 70% of actin-binding protein remains in the detergent-insoluble cell residue. The soluble actin is not filamentous as determined by sedimentation analysis, the DNAase I inhibition assay, and electron microscopy, indicating that the cytoskeleton is not fragmented by detergent extraction. The spacing between the ramifications of the actin network is 94 +/- 47 nm and 118 +/- 72 nm in cytoskeletons prepared for electron microscopy by freeze drying and critical point drying, respectively. Free filament ends are rare, except for a few which project upward from the body of the network or which extend down to the substrate. Filaments of the network intersect predominantly at right angles to form either T-shaped and X-shaped overlaps having striking perpendicularity or else Y-shaped intersections composed of filaments intersecting at 120-130 degrees angles. The actin filament concentration in the lamellae is high, with an average value of 12.5 mg/ml. The concentration was much more uniform in freeze-dried preparations than in critical point-dried specimens, indicating that there is less collapse associated with the freezing technique. The orthogonal actin network of the macrophage cortical cytoplasm resembles actin gels made with actin-binding protein. Reaction of cell cytoskeletons and of an actin gel made with actin-binding protein with anti-actin-binding protein IgG and anti-IgG-coated gold beads resulted in the deposition of clusters of gold at points where filaments intersect and at the ends of filaments that may have been in contact with the membrane before its removal with detergent. In the actin gel made with actin-binding protein, 75% of actin-fiber intersections labeled, and the filament spacing between intersections is consistent with that predicted on theoretical grounds if each added actin-binding protein molecule cross-links two filaments to form an intersection in the gel.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Ultrastructural analysis of the dynactin complex: an actin-related protein is a component of a filament that resembles F-actin.

1994 ◽  
Vol 126 (2) ◽  
pp. 403-412 ◽  
Author(s):  
D A Schafer ◽  
S R Gill ◽  
J A Cooper ◽  
J E Heuser ◽  
T A Schroer
Keyword(s):  
Electron Microscopy ◽  
Binding Protein ◽  
Terminal Region ◽  
Ultrastructural Analysis ◽  
Actin Binding ◽  
Related Protein ◽  
Actin Binding Protein ◽  
Capping Protein ◽  
Short Filament ◽  
Dynactin Complex

The dynactin complex visualized by deepetch electron microscopy appears as a short filament 37-nm in length, which resembles F-actin, plus a thinner, laterally oriented filament that terminates in two globular heads. The locations of several of the constituent polypeptides were identified on this structure by applying antibodies to decorate the dynactin complex before processing for electron microscopy. Antibodies to the actin-related protein Arp1 (previously referred to as actin-RPV), bound at various sites along the filament, demonstrating that this protein assembles in a polymer similar to conventional actin. Antibodies to the barbed-end actin-binding protein, capping protein, bound to one end of the filament. Thus, an actin-binding protein that binds conventional actin may also bind to Arp1 to regulate its polymerization. Antibodies to the 62-kD component of the dynactin complex also bound to one end of the filament. An antibody that binds the COOH-terminal region of the 160/150-kD dynactin polypeptides bound to the globular domains at the end of the thin lateral filament, suggesting that the dynactin polypeptide comprises at least part of the sidearm structure.

scholarly journals A calcium- and pH-regulated protein from Dictyostelium discoideum that cross-links actin filaments.

1982 ◽  
Vol 94 (2) ◽  
pp. 466-471 ◽  
Author(s):  
J Condeelis ◽  
M Vahey
Keyword(s):  
Electron Microscopy ◽  
Dictyostelium Discoideum ◽  
Actin Filaments ◽  
Binding Protein ◽  
Actin Binding ◽  
Cross Linking ◽  
Optimal Conditions ◽  
Maximal Inhibition ◽  
Actin Binding Protein ◽  
Cross Links

We have purified an actin binding protein from amebas of Dictyostelium discoideum which we call 95,000-dalton protein (95K). This protein is rod shaped, approximately 40 nm long in the electron microscope, contains two subunits measuring 95,000 daltons each, and cross-links actin filaments. Cross-linking activity was demonstrated by using falling-ball viscometry, Ostwald viscometry, and electron microscopy. Cross-linking activity is optimal at 0.1 microM Ca++ and pH 6.8, but is progressively inhibited at higher Ca++ and pH levels over a physiological range. Half-maximal inhibition occurs at 1.6 microM free Ca++ and pH 7.3, respectively. Sedimentation experiments demonstrate that elevated Ca++ and pH inhibit the binding of 95K to F-actin which explains the loss of cross-linking activity. Electron microscopy demonstrates that under optimal conditions for cross-linking, 95K protein bundles actin filaments and that this bundling is inhibited by microM Ca++. Severing of actin filaments by 95K was not observed in any of the various assays under any of the solution conditions used. Hence, 95K protein is a rod-shaped, dimeric, Ca++- and pH-regulated actin binding protein that cross-links but does not sever actin filaments.

Clathrin Hub Expression Dissociates the Actin-Binding Protein Hip1R from Coated Pits and Disrupts Their Alignment with the Actin Cytoskeleton

Traffic ◽  
2001 ◽  
Vol 2 (11) ◽  
pp. 851-858 ◽  
Author(s):  
Elizabeth M. Bennett ◽  
Chih-Ying Chen ◽  
Asa E. Y. Engqvist-Goldstein ◽  
David G. Drubin ◽  
Frances M. Brodsky
Keyword(s):  
Actin Cytoskeleton ◽  
Binding Protein ◽  
Actin Binding ◽  
Coated Pits ◽  
Actin Binding Protein

Binding of Human Platelet Glycoprotein lb and Actin to Fragments of Actin-Binding Protein

1992 ◽  
Vol 67 (02) ◽  
pp. 252-257 ◽  
Author(s):  
Anne M Aakhus ◽  
J Michael Wilkinson ◽  
Nils Olav Solum
Keyword(s):  
Actin Filaments ◽  
High Speed ◽  
Binding Protein ◽  
Protein A ◽  
Actin Binding ◽  
Platelet Glycoprotein ◽  
Actin Binding Protein ◽  
Crossed Immunoelectrophoresis ◽  
Triton X ◽  
Calpain Inhibitors

SummaryActin-binding protein (ABP) is degraded into fragments of 190 and 90 kDa by calpain. A monoclonal antibody (MAb TI10) against the 90 kDa fragment of ABP coprecipitated with the glycoprotein lb (GP lb) peak observed on crossed immunoelectrophoresis of Triton X-100 extracts of platelets prepared without calpain inhibitors. MAb PM6/317 against the 190 kDa fragment was not coprecipitated with the GP lb peak under such conditions. The 90 kDa fragment was adsorbed on protein A agarose from extracts that had been preincubated with antibodies to GP lb. This supports the idea that the GP Ib-ABP interaction resides in the 90 kDa region of ABP. GP lb was sedimented with the Triton-insoluble actin filaments in trace amounts only, and only after high speed centrifugation (100,000 × g, 3 h). Both the 190 kDa and the 90 kDa fragments of ABP were sedimented with the Triton-insoluble actin filaments.

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