The human erythrocyte membrane skeleton may be an ionic gel

Biochemical and biophysical observations indicate that the erythrocyte membrane skeleton is composed of a swollen network of long, flexible and ionizable macromolecules located at the cytoplasmic surface of the fluid membrane lipid bilayer. We have analyzed the mechanochemical properties of the erythrocyte membrane assuming that the membrane skeleton constitutes an ionic gel (swollen ionic elastomer). Using recently established statistical thermodynamic theory for such gels, our analysis yields mathematical expressions for the mechanochemical properties of erythrocyte membranes that incorporate membrane molecular parameters to an extent not achieved previously. The erythrocyte membrane elastic shear modulus and maximum elastic extension ratio predicted by our membrane model are in quantitative agreement with reported values for these parameters. The gel theory predicts further that the membrane skeleton modulus of area compression, K _G, may be small as well as large relative to the membrane elastic shear modulus, G, depending on the environmental conditions. Our analysis shows that the ratio between these two parameters affects both the geometry and the stability of the favoured cell shapes.     

Long-term storage of xanthan in seawater at elevated temperature: physical dimensions and chemical composition of degradation products

Commercial xanthan and xanthan from Xanthomonas strain 646 produced in the laboratory have been subjected to heat treatment for various periods of time in oxygen depleted, high salinity, aqueous solutions. Both the viscosity and the carbohydrate content decreased with increasing incubation time at a specified temperature. The losses increased with increasing temperature. Data from electron micrographs and dialysable sugar content indicate that random cleavage of the double-stranded xanthan chain is the main mechanism responsible for the decreasing viscosity. Removal of pyruvate and acetate substituents on the side chains was apparently not related to the change in physical dimensions. The mannose/glucose ratio in the non-diffusible fraction decreased with incubation time, apparently not related to change in physical dimensions. Electron micrographs showed that one of the samples appeared as highly aggregated in the native condition. After 1 month at 80 degrees C, we observed that the aggregates had dissolved and that the viscosity had increased fivefold. This suggests that heat treatment can be used to avoid microgels and to obtain higher viscosifying power of the native xanthan.     

Controlled gelation of xanthan by trivalent chronic ions

Addition of trivalent chromic ions to xanthan solutions gives rise to gel formation. The dynamic shear storage and loss moduli (0.01 – 10 rad/s) of xanthan solutions with polymer concentrations ranging from 1 to 7 mg/ml and Cr³⁺ concentrations ranging from 0 to 50 m have been studied. It is found that the rate of gel formation is strongly dependent on the Cr³⁺ concentration, but to a much smaller extent on the xanthan concentration. The gelation time is less than 1 h for 50 m Cr³⁺ and about 40 h for 2 m Cr³⁺. It is found that the minimum Cr³⁺ concentration needed to give gelation of 1–7 mg/ml xanthan is 1–2 m .     

Mode of action of recombinant Azotobacter vinelandii mannuronan C-5 epimerases AlgE2 and AlgE4.

The enzymes mannuronan C-5 epimerases catalyze conversion of beta-D-mannuronic acid to alpha-L-guluronic acid in alginates at the polymer level and thereby introduce sequences that have functional properties relevant to gelation. The enzymatic conversion by recombinant mannuronan C-5 epimerases AlgE4 and AlgE2 on alginate type substrates with different degree of polymerization and initial low fraction of alpha-L-guluronic acid was investigated. Essentially no enzymatic activity was found for fractionated mannuronan oligomer substrates with an average degree of polymerization, DP(n), less than or equal 6, whereas increasing the DP(n) yielded increased epimerization activity. This indicates that these enzymes have an active site consisting of binding domains for consecutive residues that requires interaction with 7 or more consecutive residues to show enzymatic activity. The experimentally determined kinetics of the reaction, and the residue sequence arrangement introduced by the epimerization, were modeled using Monte Carlo simulation accounting for the various competing intrachain substrates and assuming either a processive mode of action or preferred attack. The comparison between experimental data and simulation results suggests that epimerization by AlgE4 is best described by a processive mode of action, whereas the mode of action of AlgE2 appears to be more difficult to determine.     

HeLa cell plasma membranes. Changes in membrane protein composition during the cell cycle.

HeLa cells grown in suspension culture were synchronized by amethopterin block and thymidine reversal. In some cases an additional Colcemid block was used to obtain mitotic cells. From the various phases of the cell cycle, cells were harvested and the plasma membranes isolated. The membrane proteins were solubilized in sodium dodecyl sulphate and separated by gel electrophoresis in the presence of sodium dodecyl sarcosinate. About 35 protein bands, five of which were stained with periodic acid-Schiff reagent, appeared. Most of the bands were identical in all membrane preparations, but a few minor bands seemed to be associated with limited periods of the cell cycle. In particular, the cells in mitosis apparently contained plasma membrane proteins which did not occur in other phases. Amino acid analyses of the plasma membranes revealed no significant cell cycle-dependent changes in the amino acid composition.     

Thermal stability and biochemical properties of isocitrate dehydrogenase from the thermoacidophilic archaeon <Emphasis Type="Italic">Thermoplasma acidophilum</Emphasis>

Isocitrate dehydrogenase [IDH; EC 1.1.1.42] from the thermoacidophilic archaeon Thermoplasma acidophilum (TaIDH) showed high thermal stability with an apparent melting temperature, T _m, of 82.2 and 84.5°C at pH 7.5 and 5.8, respectively. Based on structural alignment of TaIDH with IDH from Aeropyrum pernix (ApIDH) and Archaeoglobus fulgidus (AfIDH) residues forming an aromatic cluster in the clasp-domain thought to strengthen the dimer interface in ApIDH and AfIDH were identified in the former enzyme. Moreover, TaIDH had a shortened N-terminus that may protect the enzyme from thermal denaturation. The enzyme activity of TaIDH was highest at 70°C. The pH-activity profile was bell-shaped with an optimum shifted to a lower pH compared to AfIDH. The activity of TaIDH was influenced by changes in pH with a three-fold reduction in activity when the pH was shifted from the pH-optimum at 7.5 to pH 5.8. However, the specific activity at pH 5.8 was still high when compared with AfIDH. The reduction in activity at pH 5.8 was not due to instability of the enzyme as the T _m of TaIDH was higher at pH 5.8 than at 7.5 and the enzyme retained 91% of its activity after incubation at 1 h at pH 5 and 60°C. The difference in the pH-profile of TaIDH in comparison with AfIDH may thus be related to the pK _as of their catalytic residues involved in the initial proton abstraction and the final proton donation during the catalysis of oxidative decarboxylation of isocitrate to 2-oxoglutarate and reduced coenzyme.