Theoretical investigations on circular and chain-like hydrogen bonded structures found in two crystal forms of α-cyclodextrin hexahydrate: Models for hydration and water clusters

Hydration of macromolecules and the structure of water of crystallization are not understood in detail because in these complex systems. H-atoms cannot be located and the hydrogen bonding schemes are not known. X-ray and neutron diffraction studies on a hydrated oligosaccharide, α-cyclodextrin 6H₂O, ((C₆H₁₀O₅)₆·6H₂O), crystals forms A and B, gave insight into the chain-like and circular arrangement of hydrogen bonds. In the circles, homodromic (unidirectional) and antidromic (counter-running) orientation of five to six hydrogen bounds is observed. PCILO calculations showed that homodromic circles and chains are approx. 8% per hydrogen bond more stable than antidromic circles, that the changes in electronic charges on H and O atoms are greater in homo than in antidromic systems and that the dipole moments are only approx. 3 D in the homodromic circles but 6–8 D in chain-like and antidromic arrangement. These results have been interpreted in terms of cooperative effect. Circular systems are considered as structural elements in hydration shells of macromolecules and in the assembly of ‘flickering’ water clusters.     

Light-dependent conformational change at rhodopsin's cytoplasmic surface detected by increased susceptibility to proteolysis

Rhodopsin in bovine photoreceptor disk membranes was subjected to limited proteolysis by thermolysin, removing twelve amino acids from rhodopsin's carboxyl terminus. (1) The rate of proteolysis is significantly faster with rhodopsin following exposure to light than with unbleached rhodopsin, provided that the incubation conditions (pH, temperature) favor the formation of metarhodopsin II. (2) If the disk membranes are illuminated under conditions in which metarhodopsin I is the predominant photoproduct (pH 8.5, 0°C), no increase in the rate of proteolysis is observed compared to unilluminated membranes. (3) The light-induced increase in the rate of proteolysis is transient: it slowly decays in the dark to the original rate found for unbleached rhodopsin. The enhanced susceptibility to proteolysis appears to measure a conformational change at rhodopsin's cytoplasmic surface which is first exhibited at the metarhodopsin II stage. This and possibly other light-dependent changes may allow rhodopsin to mediate its signal as a light-receptor protein by binding to and activating certain rod cell enzymes.     

Restoration of membrane potential in mitochondria deenergized with carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP)

The membrane potential (Δψ) of rat liver mitochondria dropped upon addition of carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) but was gradually and fully restored to the original value by the subsequent addition of dithioerythritol. Concomitantly, Ca²⁺ released from mitochondria was reaccumulated and the oxidative phosphorylation process completely recoupled. Neither of these effects has been observed with dinitro-o-cresol or 2,4-dinitrophenol, uncouplers which, unlike FCCP, do not react with thiols. Δψ abolished by FCCP was also restored, though incompletely, by albumin; a prompt and complete restoration was however achieved upon subsequent addition of dithioerythritol. Dithioerythritol also completely and rapidly restored the Δψ decreased by addition of diazene dicarboxylic acid bisdimethylamide (diamide).     

Prolactin modifies the prostaglandin synthesis, prolactin binding and fluidity of mouse liver membranes

The objective of these studies was to determine if prolactin, known to induce its own receptors, alters the prostaglandin (PG) synthesis which could, in turn, modify the fluidity of the membrane and thus alter the functionality of prolactin receptors. Adult male C₃H mice were injected subcutaneously with 100 μg of oPRL every 4 h for 0, 24 or 48 h and sacrificed 8 h after receiving the last injection. Liver 100,000 × $\text{g}$ membrane pellets were used in the measurement of these parameters. The amount of binding of prolactin to these membranes increased with the duration of injections, the values being 179 and 244% of control values after 24 and 48 h of injections, respectively. The amounts of PGF_2α and PGE synthesized also increased after these injections, the values being 127 and 270% of control for PGF_2α and 634 and 695% of control values for PGE after 24 and 48 h of injections, respectively. Fluorescence polarization, an index of microviscosity, was decreased by 14 and 20% after 24 and 48 h of PRL administration, respectively. Previous studies have demonstrated simultaneous in vitro effects of prostaglandin on both prolactin receptors and membrane fluidity. The current data are in agreement with those observations and suggest that prolactin may modulate its own receptor by increasing the fluidity of the membrane in which it exists by alterations within the PG cascade. Such biochemical changes may then modify existing restraints and allow the hormone receptor to assume a more functional configuration.     

Neonatally induced tolerance to soluble protein antigens. I. Analysis of B-cell and T helper cell function in mice tolerant to bovine serum albumin or fowl gamma-globulin

High dose tolerance to either bovine serum albumin (BSA) or fowl γ-globulin (FGG) was induced in CBA mice by neonatal injection. Tolerance to BSA lasted about 9 weeks, and that to FGG, about 18 weeks. Splenic B-cell function was analyzed using quantitative in vivo assays and in vitro limiting dilution analysis. Tolerogen-specific IgM- and non-IgM-producing B cells are depleted at least threefold in the spleens of tolerant mice. Tolerogen-specific T-helper-cell function was examined by immunization with haptenated antigens. Analysis of the recovery from tolerance indicates that the return to normal function in the tolerogen-specific B-cell and T helper fractions coincides with the return to normal responsiveness by the whole animal.     

Application of antimony microelectrodes to intracellular pH monitoring

Some novel studies of the properties of the antimony microelectrode used for intracellular pH measurements are described. First, it is shown that currents in the picoampere range, such as those encountered as leakage in some electrometers, induce important changes in pH sensitivity. The response time of the electrode has also been measured and indicates that the electrode exhibits a rapid time course which would be very useful for dynamic cytoplasmic pH investigations. An example of internal pH recording during cellular acidification in Xenopus laevis oocyte is also presented.     

Two Possibly Distinct Prostaglandin E1 Receptors in N1E-115 Clone: One Mediating Inositol Trisphosphate Formation, Cyclic GMP Formation, and Intracellular Calcium Mobilization and the Other Mediating Cyclic AMP Formation

Prostaglandin E1 (PGE1)-mediated transmembrane signal control systems were investigated in intact murine neuroblastoma cells (clone N1E-115). PGE1 increased intracellular levels of total inositol phosphates (IP), cyclic GMP, cyclic AMP, and calcium ([Ca2+]i). PGE1 transiently increased inositol 1,4,5-trisphosphate formation, peaking at 20 s. There was more than a 10-fold difference between the ED50 for PGE1 at cyclic AMP formation (70 nM) and its ED50 values at IP accumulation (1 microM), cyclic GMP formation (2 microM), and [Ca2+]i increase (5 microM). PGE1-mediated IP accumulation, cyclic GMP formation, and [Ca2+]i increase depended on both the concentration of PGE1 and extracellular calcium ions. PGE1 had more potent intrinsic activity in cyclic AMP formation, IP accumulation, and cyclic GMP formation than did PGE2, PGF2 alpha, or PGD2. A protein kinase C activator, 4 beta-phorbol 12 beta-myristate 13 alpha-acetate, had opposite effects on PGE1-mediated IP release and cyclic GMP formation (inhibitory) and cyclic AMP formation (stimulatory). These data suggest that there may be subtypes of the PGE1 receptor in this clone: a high-affinity receptor mediating cyclic AMP formation, and a low-affinity receptor mediating IP accumulation, cyclic GMP formation, and intracellular calcium mobilization.