Electron paramagnetic resonance of nitrosylprotoheme dimethyl ester complexes with imidazole derivatives as model systems for nitrosylhemoproteins.

For the purpose of understanding the electron paramagnetic resonance (epr) spectral change of nitrosylhemoproteins under various conditions, the epr spectra for the model system have been analyzed. The model system consists of the nitrogen oxide complex of the iron(II) protoporphyrin IX dimethyl ester and various imidazole derivatives (three hindered and six unhindered imidazole derivatives). The results of the analysis indicate the existence of two molecular species in the model system, which differ in structure of the FeNO unit. These observations were compared with those for the nitrosylhemoproteins.     

The age distribution of neutralizing antibodies against varicella-zoster virus in healthy individuals

A seroepidemiological survey of varicella was made in the Nagoya area by the neutralization (NT) test. Of 1,473 recorded cases of varicella, 81.4% were under 6 years old and 9.6% were under one year old; of the 168 recorded cases under one year old, about 30% were under 5 months old. Examination of 11 pairs of mother and cord sera and 13 pairs of mother and infant sera showed that transfer of NT antibody was in general good, even in babies that were small for their age or smnall at birth after 28 weeks gestation. The transferred maternal antibody decreased rapidly, becoming undetectable in babies of 4 months old. Then with increase in age the percentgage of seropositive children gradually increased, being 53.3% at 4 to 5 years old, and 100% in those of over 9 years old, with a temporary decrease in young adults in their twenties.     

Comparative studies on the inhibitory effects of interferon on various strains of herpes simplex viruses in vitro

The sensitivities to human leukocyte interferon of 10 strains of Herpes simplex virus (HSV) type 1 and 3 strains of type 2 were compared. All the strains were sensitive to interferon, although their sensitivities were less than that of vesicular stomatitis virus (VSV). There was no significant difference in the sensitivities to interferon of HSV type 1 and type-2 or among different strains of a given type of HSV. Nor was there any difference between the sensitivities of 5-iodo-2'-deoxyuridine (IDU)-sensitive and resistant strains isolated from the same patients. These results suggest that interferon should be useful in therapy of HSV infection.     

Sexual difference and organ specificity of the effect of estradiol on carbonic anhydrase and Mg(2+)-HCO3(-)-ATPase activities isolated from duodenal mucosa and kidney cortex of male and female rats: preliminary study with crude enzyme samples.

Effects of the s.c. administration of various doses of estradiol propionate (E.P.; 25-500 micrograms/kg) on the activities of carbonic anhydrase (CA), Mg(2+)-dependent ATPase and Mg(2+)-dependent, HCO3(-)-stimulated ATPase (Mg(2+)-HCO3(-)-ATPase) in rat duodenal mucosa and kidney cortex, and on body weight, organ weight and serum concentrations of testosterone and estradiol-17 beta, were examined in adult male, female, testectomized and ovariectomized rats. In normal male rats, activities of cytosol CA and brush border Mg(2+)-HCO3(-)-ATPase in the kidney were increased in a dose-dependent manner and reached 1.6- and 2-fold of controls, respectively, after consecutive administration (daily for 7 days) of 500 micrograms E.P. with no changes in either enzyme activities in duodenal mucosa. The positive correlations (P less than 0.01) were observed by linear regression analysis between serum concentration of estradiol-17 beta and kidney cytosol CA or kidney brush border Mg(2+)-HCO3(-)-ATPase activities. In normal female rats, activities of cytosol CA and brush border Mg(2+)-HCO3(-)-ATPase in the duodenal mucosa, and brush border Mg(2+)-HCO3(-)-ATPase activity in the kidney were increased by E.P. administration (100 and 500 micrograms/kg, daily for 7 days), however, kidney cytosol CA activity did not change by any dosage. Behavior of a part of both enzymes to E.P. in testectomized rats was altered almost in the same way to that observed in normal female rats and vice versa in ovariectomized rats. Body weight was decreased, in general, by consecutive administration of E.P. in a dose-dependent manner, and kidney weight was increased by E.P. in both male and female rats.     

Small-angle x-ray scattering study of metal ion-induced conformational changes in Serratia protease.

Metal ion-induced conformational changes in Serratia protease which contains one zinc ion per molecule were investigated by the small-angle x-ray scattering method. The molecule is an elongated ellipsoid of approximately 110 x 40 x 40 A with a large cleft in its central region. Comparisons of the native (zinc-enzyme) with the zinc-free (apoenzyme) enzyme and with the zinc-replated metalloenzyme show small but significant differences in their radii of gyration, maximum particle dimensions, and intraparticle pair-distance distributions. The radius of gyration and maximum particle dimension of the native enzyme are almost the same as those of the cobalt-enzyme but are shorter and longer, respectively, than those of the apo- and cadmium-enzymes. Simulation analysis based on the intraparticle pair-distribution function showed that these modified enzymes are comparable with the native enzyme in overall structure, and, except for the cobalt-enzyme, differ in cleft size. The residual enzymatic activity of the cobalt-enzyme is the same as that of the native enzyme, but the apo- and cadmium-enzymes have considerably less activity. The size of the cleft therefore is strictly controlled to ensure optimal enzyme activity, and the position and coordination behavior of the zinc ion in the cleft appears to be essential both for biological functioning and for the maintenance of the gross tertiary structure.     

Putative inositol 1,4,5-trisphosphate binding proteins in rat brain cytosol.

In previous works, we synthesized a series of inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) analogs, with a substituent on the second carbon of the inositol ring. Using these analogs, the Ins(1,4,5)P3 affinity media were also synthesized (Hirata, M., Watanabe, Y., Ishimatsu, T., Yanaga, F., Koga, T., and Ozaki, S. (1990) Biochem. Biophys. Res. Commun. 168, 379-386). When the cytosol fraction from the rat brain was applied to an Ins(1,4,5)P3 affinity column, an eluate with a 2 M NaCl solution was found to have remarkable Ins(1,4,5)P3-binding activity. The active fraction was further fractionated with gel filtration chromatography, and two proteins with an apparent molecular mass of 130 or 85 kDa were found to be Ins(1,4,5)P3-binding proteins but with no Ins(1,4,5)P3 metabolizing activities. Partial amino acid sequences determined after proteolysis and reversed-phase chromatography revealed that the protein with an apparent molecular mass of 85 kDa is the delta-isozyme of phospholipase C and that of 130 kDa has no sequence the same as the Ins(1,4,5)P3-recognizing proteins hitherto examined. Ins(1,4,5)P3 at concentrations greater than 1 microM strongly inhibited 85-kDa phospholipase C delta activity, without changing its dependence on the concentrations of free Ca2+ and H+. Among inositol phosphates examined, Ins(3,4,5,6)P4 inhibited the binding of [3H]Ins(1,4,5)P3 to the 130-kDa protein at much the same concentrations as seen with Ins(1,4,5)P3. This report seems to be the first evidence for the presence of soluble Ins(1,4,5)P3-binding proteins in the rat brain, one of which is the delta isozyme of phospholipase C.     

Evaluation of platelet calcium ion mobilization by the use of various divalent ions.

Divalent ion mobilization in human platelets was evaluated with Fura-2 fluorescence changes induced by Ca2+, Sr2+, Ba2+ and Mn2+. Extracellular Ca2+, Sr2+ and Ba2+ all entered thrombin-stimulated platelets. These divalent ions were also able to refill the intracellular Ca2+ storage sites which had been depleted of Ca2+ by ionomycin treatment, and were released from the storage sites upon thrombin stimulation. However, only the refilling of the storage sites with Ca2+ and Sr2+, but not with Ba2+, were capable of suppressing the opening state of Ca2+ channels assessed with Mn2+ influx. Efflux of intracellularly accumulated divalent ions was observed with Ca2+ and Sr2+ but not with Ba2+. These findings indicate that there are subtle differences in the Ca(2+)-binding domains of the various systems involved in Ca2+ mobilization in platelets, some of which discriminate Ba2+ while accepting Sr2+.     

Fungal coil formation ofRhizoctonia repens in seedlings ofGaleola septentrionalis (orchidaceae)

Protocorms or protocorms with roots of an achlorophyllous orchidGaleola septentrionalis were inoculated with isolates ofRhizoctonia repens, R. solani, andRhizoctonia spp. The seedlings were infected with eight of twelve isolates ofR. repens. Fungal coils were formed in the cells, which was suggestive of a symbiotic association. The other isolates caused soft rot or no infection to the protocorms or the protocorms with a root. Contribution No. 97, Laboratories of Plant Pathology and Mycology, Institute of Agriculture and Forestry, University of Tsukuba, Tsukuba, Ibaraki 305, Japan.     

Molecular cloning and nucleotide sequencing of the gene for E. coli cAMP receptor protein.

The crp gene of E. coli, which codes for cAMP receptor protein (CRP), has been cloned in the plasmid pBR322 on the basis of a genetic complementation. One of the recombinant plasmids, pHA1, was shown to direct the synthesis of CRP in a cell-free system. The location of the crp gene was determined by constructing subclones carrying various portions of pHA1. The nucleotide sequence of the crp gene has been determined. The coding region consists of 627 base pairs (bp), which specify a protein of 209 amino acids. The predicted amino acid sequence from the DNA sequence is consistent with the amino acid sequence partially known and the amino acid composition of CRP. After the coding region, there is a G-C rich inverted repeat sequence followed by a run of Ts, which could be a terminator of the crp gene. A possible promoter sequence was found about 180 bp upstream from the initiation codon and was shown to act as a promoter in vitro and in vivo. There are two dyad symmetry regions in a 167 bp leader sequence.