scholarly journals Isolation and characterization of two forms of a cytoskeleton.

1979 ◽  
Vol 83 (1) ◽  
pp. 109-115 ◽  
Author(s):  
K T Edds
Keyword(s):  
Sea Urchin ◽  
Electron Micrographs ◽  
Actin Filaments ◽  
Three Dimensional ◽  
Strongylocentrotus Droebachiensis ◽  
Major Protein ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Filament Bundles

Isolated petaloid coelomocytes from the sea urchin Strongylocentrotus droebachiensis transform to a filopodial morphology in hypotonic media. Electron micrographs of negatively stained Triton-insoluble cytoskeletons show that the petaloid form consists of a loose net of microfilaments while the filopodial form consists of paracrystalline bundles of microfilaments. Actin is the major protein of both forms of the cytoskeleton. Additional polypeptides have molecular weights of approximately 220,000, 64,000, 57,000, and 27,000 daltons. Relative to actin the filopodial cytoskeletons have an average of 2.5 times as much 57k polypeptide as the petaloid cytoskeletons. Treatment with 0.25 M NaCl dissociates the filament bundles into individual actin filaments free of the actin-associated polypeptides. Thus, one or more of these actin-associated polypeptides may be responsible for crosslinking the actin filaments into bundles and maintaining the three-dimensional nature of the cytoskeletons.

scholarly journals α-Crystallin. The isolation and characterization of distinct macromolecular fractions

1971 ◽  
Vol 124 (2) ◽  
pp. 337-343 ◽  
Author(s):  
Abraham Spector ◽  
Lu-Ku Li ◽  
Robert C. Augusteyn ◽  
Arthur Schneider ◽  
Thomas Freund
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Chemical Difference ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Different Populations ◽  
Molecular Weight Fractions

α-Crystallin was isolated from calf lens periphery by chromatography on DEAE-cellulose and gel filtration. Three distinct populations of macromolecules have been isolated with molecular weights in the ranges approx. 6×105−9×105, 0.9×106−4×106and greater than 10×106. The concentration of macromolecules at the molecular-weight limits of a population are very low. The members of the different populations do not appear to be in equilibrium with each other. Further, in those molecular-weight fractions investigated, no equilibrium between members of the same population was observed. The population of lowest molecular weight comprises 65–75% of the total material. The amino acid and subunit composition of the different-sized fractions appear very similar, if not identical. The only chemical difference observed between the fractions is the presence of significant amounts of sugar in the higher-molecular-weight fractions. Subunit molecular weights of approx. 19.5×103and 22.5×103were observed for all α-crystallin fractions.

scholarly journals Isolation and Characterization of Mutations in theEscherichia coli Regulatory Protein XapR

1999 ◽  
Vol 181 (14) ◽  
pp. 4397-4403 ◽  
Author(s):  
Casper Jørgensen ◽  
Gert Dandanell
Keyword(s):  
Amino Acids ◽  
Purine Nucleoside Phosphorylase ◽  
Mutational Analysis ◽  
Regulatory Protein ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Wild Type ◽  
Isolation And Characterization ◽  
In The Wild

ABSTRACT In this work, the LysR-type protein XapR has been subjected to a mutational analysis. XapR regulates the expression of xanthosine phosphorylase (XapA), a purine nucleoside phosphorylase inEscherichia coli. In the wild type, full expression of XapA requires both a functional XapR protein and the inducer xanthosine. Here we show that deoxyinosine can also function as an inducer in the wild type, although not to the same extent as xanthosine. We have isolated and characterized in detail the mutants that can be induced by other nucleosides as well as xanthosine. Sequencing of the mutants has revealed that two regions in XapR are important for correct interactions between the inducer and XapR. One region is defined by amino acids 104 and 132, and the other region, containing most of the isolated mutations, is found between amino acids 203 and 210. These regions, when modelled into the three-dimensional structure of CysB from Klebsiella aerogenes, are placed close together and are most probably directly involved in binding the inducer xanthosine.

scholarly journals Isolation and Characterization of 2,3-Dichloro-1-Propanol-Degrading Rhizobia

2000 ◽  
Vol 66 (7) ◽  
pp. 2882-2887 ◽  
Author(s):  
Agus J. Effendi ◽  
Steven D. Greenaway ◽  
Brian N. Dancer
Keyword(s):  
Molecular Weight ◽  
High Ph ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Complete Mineralization ◽  
Native Molecular Weight

ABSTRACT 2,3-Dichloro-1-propanol is more chemically stable than its isomer, 1,3-dichloro-2-propanol, and is therefore more difficult to degrade. The isolation of bacteria capable of complete mineralization of 2,3-dichloro-1-propanol was successful only from enrichments at high pH. The bacteria thus isolated were found to be members of the α division of the Proteobacteria in the Rhizobiumsubdivision, most likely Agrobacterium sp. They could utilize both dihaloalcohol substrates and 2-chloropropionic acid. The growth of these strains in the presence of 2,3-dichloro-1-propanol was strongly affected by the pH and buffer strength of the medium. Under certain conditions, a ladder of four active dehalogenase bands could be visualized from this strain in activity gels. The enzyme involved in the complete mineralization of 2,3-dichloro-1-propanol was shown to have a native molecular weight of 114,000 and consisted of four subunits of similar molecular weights.

Isolation and Characterization of Low Density Detergent-Insoluble Membrane (LD-DIM) Fraction from Sea Urchin Sperm

1999 ◽  
Vol 258 (3) ◽  
pp. 616-623 ◽  
Author(s):  
Kaoru Ohta ◽  
Chihiro Sato ◽  
Tsukasa Matsuda ◽  
Masaru Toriyama ◽  
William J. Lennarz ◽  
...  
Keyword(s):  
Sea Urchin ◽  
Low Density ◽  
Isolation And Characterization ◽  
Sea Urchin Sperm

scholarly journals Structure, Subunit Topology, and Actin-binding Activity of the Arp2/3 Complex from Acanthamoeba

1997 ◽  
Vol 136 (2) ◽  
pp. 331-343 ◽  
Author(s):  
R. Dyche Mullins ◽  
Walter F. Stafford ◽  
Thomas D. Pollard
Keyword(s):  
Electron Micrographs ◽  
Actin Filaments ◽  
Analytical Ultracentrifugation ◽  
Fluorescent Antibody ◽  
Complex Molecule ◽  
Gel Filtration ◽  
Sedimentation Coefficient ◽  
Leading Edge ◽  
Actin Binding ◽  
Filament Bundles

The Arp2/3 complex, first isolated from Acanthamoeba castellani by affinity chromatography on profilin, consists of seven polypeptides; two actinrelated proteins, Arp2 and Arp3; and five apparently novel proteins, p40, p35, p19, p18, and p14 (Machesky et al., 1994). The complex is homogeneous by hydrodynamic criteria with a Stokes' radius of 5.3 nm by gel filtration, sedimentation coefficient of 8.7 S, and molecular mass of 197 kD by analytical ultracentrifugation. The stoichiometry of the subunits is 1:1:1:1:1:1:1, indicating the purified complex contains one copy each of seven polypeptides. In electron micrographs, the complex has a bilobed or horseshoe shape with outer dimensions of ∼13 × 10 nm, and mathematical models of such a shape and size are consistent with the measured hydrodynamic properties. Chemical cross-linking with a battery of cross-linkers of different spacer arm lengths and chemical reactivities identify the following nearest neighbors within the complex: Arp2 and p40; Arp2 and p35; Arp3 and p35; Arp3 and either p18 or p19; and p19 and p14. By fluorescent antibody staining with anti-p40 and -p35, the complex is concentrated in the cortex of the ameba, especially in linear structures, possibly actin filament bundles, that lie perpendicular to the leading edge. Purified Arp2/3 complex binds actin filaments with a Kd of 2.3 μM and a stoichiometry of approximately one complex molecule per actin monomer. In electron micrographs of negatively stained samples, Arp2/3 complex decorates the sides of actin filaments. EDC/NHS cross-links actin to Arp3, p35, and a low molecular weight subunit, p19, p18, or p14. We propose structural and topological models for the Arp2/3 complex and suggest that affinity for actin filaments accounts for the localization of complex subunits to actinrich regions of Acanthamoeba.

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