Relaxation and electrophoretic effects in polyelectrolyte solutions. I. Salt-free solutions

Relations between relaxation and electrophoretic effects in salt-free polyelectrolyte solutions are studied in terms of nonequilibrium thermodynamics and of binary friction coefficients. The significance and the method of determining these coefficients are explained. Consideration of experimental results and of long- and short-range interactions suggests that the concept of “ion condensation” is needed to provide a consistent desciption of transport processes for linear flexible polyelectrolytes. Empirical relations previously used are shown to have only limited validity.     

Stability of rhodopsin in detergent solutions.

The thermal stability of lipid-free rhodopsin in solutions of a homologous series of alkyltrimethylammonium bromide detergents and one nonionic detergent, dodecyl-beta-maltoside, has been studied as a function of detergent concentration. Rhodopsin thermal stability increases with increasing chain length within the homologous series of ionic detergents, and for chain lengths greater than 10 carbon atoms increases with increasing detergent concentration up to a "critical" concentration that depends on the chain length. Stability also increases with increasing detergent concentration for rhodopsin in solutions of the nonionic detergent. These results may be rationalized in terms of the dependence of micelle packing density on the detergent chain length, head group, and concentration.     

Aggregation of tyrocidine in aqueous solutions.

The formation of aggregates of tyrocidine B at 4°C and 20°C in aqueous solutions was studied by means of light scattering and fluorescent techniques. The apparent weight molecular weight of tyrocidine B aggregates was found to be 36,000 at 4°C and 28,800 at 20°C. Fluorescence titration experiments with dansyl-chloride resulted in an aggregational number of 31 (4°) and 28 (20°) indicating that one molecule of dye is bound per monomer of molecular weight 1,200. From a Scatchard plot apparent association constants of 1.22 × 10⁵ M (4°) and 0.95 × 10⁵ M (20°) were calculated. From the angular dependence of scattered intensity the radii of gyration were determined to be 60 Å and 58 Å, respectively.     

Macromolecular diffusion in crowded solutions.

The effects of crowding on the self or tracer diffusion of macromolecules in concentrated solutions is an important but difficult problem, for which, so far, there has been no rigorous treatment. Muramatsu and Minton suggested a simple model to calculate the diffusion coefficient of a hard sphere among other hard spheres. In this treatment, scaled particle theory is used to evaluate the probability that the target volume for a step in a random walk is free of any macromolecules. We have improved this approach by using a more appropriate target volume which also allows the calculation to be extended to the diffusion of a hard sphere among hard spherocylinders. We conclude that, to the extent that proteins can be approximated as hard particles, the hindrance of globular proteins by other proteins is reduced when the background proteins aggregate (the more so the greater the decrease in particle surface area), the hindrance due to rod-shaped background particles is reduced slightly if the rod-like particles are aligned, and the anisotropy of the diffusion of soluble proteins among cytoskeletal proteins will normally be small.     

Helix propagation in trifluoroethanol solutions.

Helix propagation of the S-peptide sequence (residues 1-19 of ribonuclease A) in 2,2,2-trifluoroethanol (TFE) solutions has been investigated with CD and nmr Overhauser effect spectroscopies. In this study, the S-peptide helix is covalently initiated at the N-terminus through disulfide bonds to a helix scaffold derived from the N-terminal sequence of the bee venom peptide apamin. The entire S-peptide sequence of this hybrid sequence peptide becomes helical at high proportions of TFE. Residues 14-19 of the S-peptide are not helical in the free peptide in TFE, nor are they helical in ribonuclease A. The "helix stop" signal encoded by the S-peptide sequence near residue 13 does not persist at high TFE with this hybrid sequence peptide. The helix-stabilizing effects of TFE are due at least in part to facilitated propagation of an extant helix. This stabilizing effect appears to be a general solvation effect and not due to specific interaction of the helical peptide with TFE. Specifically these data support the idea that TFE destabilizes the coil state by less effective hydrogen bonding of the peptide amide to the solvent.     

Quantitation of liquid-crystalline ordering in F-actin solutions.

Actin filaments (F-actin) are important determinants of cellular shape and motility. These functions depend on the collective organization of numerous filaments with respect to both position and orientation in the cytoplasm. Much of the orientational organization arises spontaneously through liquid crystal formation in concentrated F-actin solutions. In studying this phenomenon, we found that solutions of purified F-actin undergo a continuous phase transition, from the isotropic state to a liquid crystalline state, when either the mean filament length or the actin concentration is increased above its respective threshold value. The phase diagram representing the threshold filament lengths and concentrations at which the phase transition occurs is consistent with that predicted by Flory's theory on solutions of noninteracting, rigid cylinders (Flory, 1956b). However, in contrast to other predictions based on this model, we found no evidence for the coexistence of isotropic and anisotropic phases. Furthermore, the phase transition proved to be temperature dependent, which suggests the existence of orientation-dependent interfilament interactions or of a temperature-dependent filament flexibility. We developed a simple method for growing undistorted fluorescent acrylodan-labeled F-actin liquid crystals; and we derived a simple theoretical treatment by which polarization-of-fluorescence measurements could be used to quantitate, for the first time, the degree of spontaneous filament ordering (nematic order parameter) in these F-actin liquid crystals. This order parameter was found to increase monotonically with both filament length and concentration. Actin liquid crystals can readily become distorted by a process known as "texturing." Zigzaging and helicoidal liquid crystalline textures which persisted in the absence of ATP were observed through the polarizing microscope. Possible texturing mechanisms are discussed.     

Dielectric relaxation of DNA in aqueous solutions.

Using a four-electrode cell and a new electronic system for direct detection of the frequency differences specturm of solution impedance, the complex dielectric constant of calf thymus DNA (M_r = 4 × 10⁶) in aqueous NaCl at 10°C is measured at frequencies ranging from 0.2 Hz to 30 kHz. The DNA concentrations are C_p = 0.01% and 0.05%, and the NaCl concentrations are varied from C_s = 10^?4 M to 10^?3 M. A single relaxation regions is found in this frequency range, the relaxation frequency being 10 Hz at C_p = 0.01% and C_s = 10^?3 M. At C_p = 0.05% it is evidenced that the DNA chains have appreciable intermolecular interactions. The dielectric relaxaton time τ_d at C_p = 0.01% agrees well with the rotational relaxation time estimated from the reduced visocisty on the assumption that the DNA is not representable as a rigid rod but a coiled chain. It is concluded that the dielectric relaxiatioinis ascribed to the rotation of the molecule. Observed values of dielectric increment and other experimental findings are reasonably explained by assuming that the dipole moment of DNA results from the slow counterion fluctuation which has a longer relaxation time than τ_d.     

Collective behaviour of hemoglobin in dense solutions.

The formation of radical pairs of spin-labelled hemoglobin molecules (beta-93 cysteine) in a mixture with unlabelled hemoglobin of concentrations higher than the critical one [(5.7 less than c less than 6.3) mM/sigma per heme] is due to some kind of long-range order characterizing these solutions. Labelled molecules induce pertubation in an unlabelled hemoglobin solution of concentration higher than the critical one. The collective behaviour of unlabelled molecules gives rise to the formation of radical pairs of spin-labelled hemoglobin molecules. Such behaviour is dependent on the ligand bound at the heme iron (carbonmonoxide, oxygen, or water.)     

Effect of salt solutions on radiosensitivity of mammalian cells. III. Treatment with hypertonic solutions.

V79 Chinese hamster cells were treated with hypertonic solutions of NaCl or KCl and irradiated rat various times before, during, or after exposure to the solution. In solutions of molarities between 0-2 and 0-5 M, the cellular radiosensitivity increases with the molarity of the bathing solution. At these molarities, the hypertonic solution need not be present during irradiation to sensitize cells. Furthermore, radiosensitivity of cells could be increased by exposing cells for longer times to the hypertonic solution before irradiation. At higher salt concentrations (at 1-5 to 1-8 M), significant radioprotection is observed. Survival curve data showed that this protection was characterized by an increase in DO and a decrease in n, while the survival curves of cells sensitized with 0-465 M NaCl or with lower concentrations exhibited mainly changes in DO. The 1-55 M NaCl solution must be present during radiation to give a protective effect. Prolonged exposure to the salt before irradiation reduced the amount of radioprotection afforded by the salt. The results are discussed in terms of the effects of ions on histones, cellular water structure and the cell-aging cycle.     

Thermodynamics of porphyrin dimerization in aqueous solutions.

The dimerization equilibrium of deuteroporphyrin IX and of mesoporphyrin IX in aqueous solutions were studied by fluorimetric techniques over the 0.01-1 microM concentration range, where dimerization is the dominant aggregation process. Deuteroporphyrin IX was studied at several temperatures over the range 22-37 degrees C, and mesoporphyrin at 25 and 37 degrees C. The magnitudes determined for the dimerization equilibrium constants (25 degrees C, neutral pH, phosphate-buffered saline) are 2.3 X 10(6)M-1 and 5.4 X 10(6)M-1 for the deutero and meso derivatives respectively. The meso, deutero and haemato species tested show a similar temperature effect, namely dimerization decreasing with increasing temperature, indicating the involvement of a negative enthalpy change. Van''t Hoff isochore of the dimerization constants determined for deuteroporphyrin IX was linear within the temperature range of 22-37 degrees C, allowing the calculation of the thermodynamic parameters. For deuteroporphyrin dimerization, those were found to be delta G0 = -36. 4kJ X mol-1; delta H0 = -46. 0kJ X mol-1 and delta S0 = -32.2J X K-1 X mol-1 (at neutral pH, 25 degrees C, phosphate-buffered saline), showing the process to be enthalpy-driven. Similar trends have been found for porphyrin species other than those studied here. Our data fit with a hypothesis giving a major role to the solvent in driving porphyrins to aggregate in aqueous solution. The magnitudes and directions of the energetic changes fit better with the expectation of the '' solvophobic force'' theory predicting enthalpy-driven association, than with the classic hydrophobic bonding, predicting the association to be entropy-driven.     

Viability of Acanthamoeba cysts in ophthalmic solutions.

Acanthamoeba keratitis is a chronic infection of the human cornea. Many people who have this infection wear soft contact lenses. Usually lens wearers clean and maintain their lenses with various ophthalmic solutions including homemade saline. Recently it has been shown that homemade saline solutions play a role in lens contamination and thus in Acanthamoeba keratitis. We therefore evaluated the viability of cysts of three species of Acanthamoeba by exposing them for various time periods to saline, cleaning, and disinfectant solutions generally used to care for these lenses. We found that the viability of the cysts in saline solutions ranged from a minimum of 14 days to 90 days of exposure. In cleaning solutions, the survival times ranged from a minimum of 1 day to 90 days of exposure. Disinfectants, as expected, were the most effective of all tested solutions in killing Acanthamoeba cysts. The survival times ranged from 6 h to 14 days. None of these products were effective in destroying Acanthamoeba cysts in less than 6 h of exposure, which exceeds the suggested time that any given solution should be used for lens care.