Glutamic acid-dihydrogen phosphate hydrogen-bonded networks: their proton polarizability as a function of cations present. Infrared investigations.

Glutamic acid [(L-glu)n] + dihydrogen phosphate systems are studied by infrared (IR) spectroscopy dried and hydrated at 75% relative humidity, as a function of both the phosphate-glutamic acid residue (Pi/glu) ratio and the type of cations present. It is shown that the glutamic acid groups form hydrogen-bonded chains with the phosphates. In these chains the positive charge fluctuates, and they show very large proton polarizability which increases in the series Li+,Na+,K+ systems. These chains are cross-linked via phosphate-phosphate hydrogen bonds, in which the proton is almost localized at one Pi. The comparison of the (L-glu)n + dihydrogen phosphate systems with the results obtained earlier in the case of (L-glu)n + hydrogen phosphate systems shows that the behavior of (L-glu)n + Pi systems strongly depends on the pH. Only with decreasing pH the conducting chains are formed. Finally, a hypothesis is discussed with regard to the charge conduction in the F0 subunit of the H+-ATPase in mitochondria.     

Stimulation of pancreatic secretory process in the rat by low-molecular weight proteinase inhibitor

Oral application of a single dose of a new synthetic proteinase inhibitor Camostate (Foy-305) in male Wistar rats was carried out together with studies of in vitro amino acid incorporation followed by separation of proteins by two-dimensional gel electrophoresis. The aim of this experiment was to analyze changes produced by the inhibitor in total protein and individual enzyme biosynthesis. Administration of 100 mg/kg Foy-305 resulted in significant inhibition of total pancreatic protein synthesis, without changes in fractional rates for individual enzymes. 50 mg/kg Foy-305 induced a 10-fold elevation of cholecystokinin (CCK) levels in serum; this persisted for 3 h and led to a significant increase in the total rate of protein synthesis with peak values at 6 and 9 h (78% and 84% above control levels, respectively), returning to control by 15 h. Changes in fractional rates of synthesis occurred with a latency of 6 h and were restricted to amylase and the anionic form of trypsinogen and chymotrypsinogen. Amylase biosynthesis decreased by about 40% from control levels at 9 h to return to control levels by 15 h. Increased synthesis of trypsinogen and chymotrypsinogen was observed; this was also phasic. The results show similar enzyme-specific regulation as previously described for exogenous CCK stimulation and for the adaptation of the pancreas to diets enriched in protein. They demonstrate the effectiveness of pulsatory endogenous hormone release in the regulation of protein synthesis.     

Mammalian DNA polymerase alpha: a replication competent holoenzyme form from calf thymus.

A complex "replication competent" holoenzyme form of DNA polymerase alpha (RC-alpha) was purified 10,000 fold from calf thymus through the use of an assay employing primed single stranded circular DNA template. The RC-alpha form could partially replicate a double-stranded oligo(dT)-tailed linear DNA and could completely convert primed single-stranded circular DNA to its double stranded form. The RC-alpha was resolved by denaturing gel electrophoresis into at least 10 discrete polypeptide species ranging in apparent molecular mass from 200 to 47 kilodaltons; three of the bands (apparent Mr of 200, 118 and 63 kilodaltons) displayed DNA polymerase activity in denaturing gel activity assay. The isolation of RC-alpha required the use of absolutely fresh calf thymus, the inclusion of ATP and protease inhibitors throughout the purification procedure. Treatment of the RC-alpha with the neutralizing anti-DNA polymerase alpha monoclonal antibody SJK 132-20 (Tanaka et al. (1982), J. Biol. Chem. 257, 8386-8390) in nondenaturing conditions selected the complete set of 10 polypeptides, whereas treatment in denaturing conditions selected the 200 kilodalton catalytic DNA polymerase active polypeptide. The properties and the behaviour of the RC-alpha preparation following removal of specific polypeptides strongly suggested that the capacity of RC-alpha to extend and replicate long template requires the function of nonproteolysed form of the 200 kilodaltons catalytic DNA polymerase core and at least 6 other auxiliary polypeptides of, respectively, 98, 87, 63, 54, 49 and 47 kilodaltons.     

Induction of specific-locus and dominant lethal mutations in male mice by chlormethine

Chlormethine (WHO), a nitrogen mustard (2,2'-dichloro-N-methyldiethylamine), induces dominant lethal and specific-locus mutations in spermatozoa and spermatids of mice.     

Histidine modulates the clastogenic effect of oxidative stress

The kinetics of the formation of single-strand breaks induced by H2O2 in DNA is more rapid in Eagle's Modified Minimal Essential Medium (MEM) than in phosphate-buffered saline (PBS). Among the components of MEM, we found that histidine increases the rate of degradation of DNA by H2O2 in PBS dose-dependently. In hamster lung fibroblasts histidine increases the cytotoxicity of H2O2, as well as the number of sister-chromatid exchanges and the frequency of micronuclei induced by hydrogen peroxide.     

Cytogenetic effects of 3,4-dichloroaniline in human lymphocytes and V79 Chinese hamster cells

3,4-Dichloroaniline (3,4-DCA), an intermediate in various chemical syntheses, has been detected as an environmental contaminant in surface waters and in the effluents from dye-manufacturing plants. Tested for clastogenicity in human lymphocytes in vitro the compound was inactive in the chromosome aberration assay yet exhibited a positive sister-chromatid exchange response in the presence of a mammalian metabolic activation system. Exposure of V79 Chinese hamster cells to 3,4-DCA caused a concentration-dependent increase in the incidence of spindle disturbances, predominantly of the initial c-mitotic type. The results indicate that 3,4-DCA might induce aneuploidy in mammalian cells by interaction with the mitotic apparatus.     

Two isozymes of dihydroxyacetone phosphate reductase in dunaliella

Two isoforms of dihydroxyacetone phosphate reductase were present in Dunaliella tertiolecta. The major form was located in the chloroplast and the minor form in the cytosol. The chloroplastic reductase eluted first from a DEAE cellulose column followed immediately by the cytosolic form. Both forms were unstable and cold labile. Addition of 5 millimolar dithiothreitol helped to stabilize the enzymes. The cytosolic isoform of DHAP reductase was detected only if the cells were in an active log phase of growth. Then its activity was 20 to 30% of the total reductase activity. When cell cultures entered late log phase of growth the activity of the cytosolic form of the enzyme disappeared, but the chloroplastic form remained. The cytosolic DHAP reductase from Dunaliella has some properties similar to the cytosolic isoform from spinach leaves. Detergents inhibited both enzymes. However, neither form of the algal dihydroxyacetone phosphate reductase was stimulated by fructose 2,6-bisphosphate. In Dunaliella the properties of the chloroplastic form were those expected for glycerol production for osmoregulation, whereas the cytosolic form, like the reductases in leaves, is more likely involved in glycerol phosphate formation for lipid synthesis.     

Radiolysis of DNA in aqueous solution in the presence of a scavenger: A kinetic model based on a nonhomogeneous reaction of OH radicals with DNA molecules of spherical or cylindrical shape

Summary Two expressions are given for the survival dose of DNA exposed to high-energy radiation in aqueous solution in the presence of a scavenger. They are derived from a model where a diffusion controlled reaction of OH radicals occurs on the surface of the DNA macromolecules in competition with scavenging in the bulk of the solution. The DNA molecules are approximated either by spheres or by cylinders. The model based on molecules of spherical shape corresponds closely to that developed by van Rijn et al. [20]. Expressions obtained from the cylindrical model are used to account for the dependence on the scavenger concentration of some experimentally measured quantities, namely the survival dose and theG value for single-strand breaks upon Co -irradiation ofX 174 DNA and polyadenylic acid, respectively.In memoriam Prof. Dr. O.E. Polansky