Synthesis of an azadioxa-planar triphenylborane and investigation of its structural and photophysical properties.

We report here the first successful synthesis of planar triphenylborane 1 with the phenyl groups bridged by oxygen and nitrogen atoms via double nucleophilic aromatic substitution reaction. The hetero atom-bridged 1 has excellent planarity. Its structural and photophysical properties are tunable by altering the bridging atoms.

Cerebral nocardia masquerading as metastatic CNS disease in an endometrial cancer patient.

Nocardia is a bacterial infection primarily originating from organic rich soil, endemic to several international geographic locations. We present the case of a 61-year-old woman previously treated for endometrial carcinoma, who three years later developed metastatic pulmonary disease and received systemic chemotherapy. After five months, she developed a large right posterior lobe lesion, suspicious for metastatic CNS disease. However, following neurosurgical resection of the lesion and infectious disease consultation, a diagnosis of nocardia was made.

Array CGH - A Powerful Tool in Molecular Diagnostic of Pathogenic Microdeletions - Williams-Beuren Syndrome - A Case Report.

Williams-Beuren syndrome (WBS) (OMIM 194050) is caused by interstitial deletions or duplications of the 7q11.23 chromosomal region and characterised through a complex phenotype. We described a case diagnosed clinically and genetically confirmed through aCGH. Genetic assessment identified three microdeletions with a total size of 1.35 Mb located at 7q11.23. The deleted regions encompasses more than 30 genes including several protein coding genes such as ELN, LIMK1, FZDS, WBSCR22, WBSCR27, WBSCR28, STX1A, CLDN3, CLDN4, LAT2, ABHD11 or EIF4H .

The reaction of sulfhydryl reagents with bovine hepatic monoamine oxidase. Evidence for the presence of two cysteine residues essential for activity.

The bovine liver monoamine oxidase (EC 1.4.3.4) was found to be inactivated by various well-known sulfhydryl reagents like p-mercuribenzoate, methylmercuric iodide and 5,5'-dithiobis-(2-nitro benzoic acid). The present investigation shows that the inactivation of the enzyme results from reactions of these reagents with 2 out of 8 titratable sulfhydryl groups per 10(5) g of the enzyme. The substrate, benzylamine, and competitive inhibitors like benzaldehyde, p-nitrobenzaldehyde, benzyl alcohol protected the enzyme from inactivation by the mercurials or the Ellman reagent. The inactivation experiments with these sulfhydryl reagents, the protection experiments, and the kinetics as well as physicochemical observations suggest that there are only two cysteine residues that are required for activity of the enzyme. It is possible that these two residues may be active-center residues.

Evidence for farnesol-mediated isoprenoid synthesis regulation in a halophilic archaeon, Haloferax volcanii.

Farnesol strongly inhibited growth of a halophilic archaeon, Haloferax volcanii, with an IC50 value of only 2 microM (0.4 microgram/ml) in rich medium and 50 nM (0.01 microgram/ml) in minimal medium without lysis. Other isoprenoid alcohols such as isopentenol, dimethylallyl alcohol, geraniol, and geranylgeraniol at 500 microM did not affect its growth. Mevalonate, which is the precursor of all isoprenoid membrane lipids in archaea, led to recovery of the growth inhibition of H. volcanii, but acetate had no such effect. Farnesol inhibited incorporation of acetate, but not mevalonate, into the lipid fraction. These results suggest that farnesol inhibited the biosynthetic pathway from acetate (acetyl-CoA) to mevalonate. Farnesol is known to be derived from the important intermediate of isoprenoids, farnesyl diphosphate (FPP), and found in neutral lipid fraction from this archaeon. Moreover, the cell-free extracts from H. volcanii could phosphorylate farnesol with ATP to generate farnesyl monophosphate and FPP. We conclude that farnesol-mediated isoprenoid synthesis regulation system by controlling farnesol concentration is present in H. volcanii.

Pressure-induced expression of heat shock protein 70 mRNA in adult rat heart is coupled both to protein kinase A-dependent and protein kinase C-dependent systems.

BACKGROUND: Production of heat shock protein 70 (HSP70) in the heart is induced by hemodynamic stress, but its intracellular signal transduction system has not been elucidated well. OBJECTIVE: To investigate the hypothesis that protein kinase A (PKA)-dependent and protein kinase C (PKC)dependent systems are involved in the pressure-induced expression of HSP70 mRNA in perfused adult rat heart METHODS: Isolated tetrodotoxin-arrested Sprague-Dawley rat hearts were perfused as Langendorff preparations at a constant aortic pressure of 60 mmHg. Aortic pressure in rats of the pressure-overloaded group was elevated from 60 to 120 mmHg for 2-120 min. cAMP contents and rates of synthesis of protein were measured by radioimmunoassay and the incorporation of [14C]-phenylalanine into total heart protein, respectively. Expression of HSP70 mRNA was determined by Northern blot analysis. RESULTS: Elevation of aortic pressure significantly increased cAMP content after 2 min of perfusion (by 41%), significantly increased rates of synthesis of protein during the second hour of perfusion (by 41%), and induced expression of HSP70 mRNA maximally after 60 min of perfusion (2.7-fold the control value). Exposure to glucagon, forskolin or 1 -methyl-3-isobutylxanthine mimicked increases in these parameters caused by elevation of aortic pressure. Administration of a selective PKA inhibitor, H-89, significantly prevented induction of increases in expression of HSP70 mRNA and rates of synthesis of protein by a high pressure overload and exposure to agents that increase cAMP content. Furthermore, administration of phorbol ester induced expression of HSP70 mRNA. Administration of a PKC inhibitor, calphostin C, significantly prevented induction of increases in expression of HSP70 mRNA by a pressure overload and by exposure to phorbol ester. CONCLUSIONS: These results suggest that the pressure-induced induction of production of HSP70 is regulated both by PKA-dependent and by PKC-dependent systems during periods of active synthesis of protein in adult rat heart.

Synthesis of peptide aldehyde derivatives as selective inhibitors of human cathepsin L and their inhibitory effect on bone resorption.

Cathepsin L, a lysosomal cysteine protease, is secreted by osteoclasts and participates in bone collagen degradation. In a search for cathepsin L inhibitors as antiosteoporotic agents, a series of peptide aldehyde derivatives were prepared by two synthetic approaches, DMSO oxidation of the corresponding alcohol derivatives and DIBAL-H reduction of the corresponding N, O-dimethylhydroxylamide derivatives, and evaluated for inhibitory activity against human cathepsin L and for inhibitory effects on bone resorption. Some of the peptide aldehyde derivatives including alpha-acylamino aldehyde derivatives showed potent activities. Among these compounds, N-(1-naphthalenylsulfonyl-L-isoleucyl-L-tryptophanal (12) was selected as a candidate for further investigation. Compound 12, a potent, selective, and reversible inhibitor of human cathepsin L with an IC50 of 1.9 nM, inhibited the release of Ca2+ and hydroxyproline from bone in in vitro bone culture system and also prevented bone loss in ovariectomized mice at an oral dose of 50 mg/kg.

Inhibition of rat liver rhodanese by di-, tricarboxylic, and alpha-keto acids.

Rat liver rhodanese [EC 2.8.1.1] purified by ammonium sulfate fractionation, CM-cellulose and Sephadex G-200 chromatography yielded two active fractions (I & II). Their molecular weights were estimated to be 1.75 X 10(4) (I) and 1.26 X 10(4) (II) by the gel filtration method. Kinetic studies revealed that Fraction I rat liver rhodanese catalyzes thiocyanate formation from thiosulfate and cyanide by a double displacement mechanism. Carboxylic acids such as DL-isocitric, citric malic, pyruvic, and oxaloacetic acid were competitive inhibitors with respect to thiosulfate, whereas fumaric, succinic, and alpha-ketoglutaric acids were noncompetitive inhibitors with respect ot thiosulfate. Incubation of mitochondria with sulfate and alpha-ketoglutaric acid caused a significant decrease in rhodanese activity.

Purification and some properties of beta-transglycosylase of Trichoderma longibrachiatum.

A beta-transglycosylase was purified to a homogeneous state from the extract of a wheat bran Koji culture of Trichoderma longibrachiatum by column chromatography. The purified enzyme showed a typical disproportionation reaction with cellopentaose as the substrate, producing a high molecular component (a water-insoluble glucan). The enzyme showed neither cellulase nor beta-glucosidase activity. The reaction was optimal at pH 6.0 and 37 degrees C. The molecular weight of the enzyme was estimated to be 11,000 by gel filtration using a TOYOPEARL HW-55F column. The amount of the glucan synthesized by the enzyme increased with prolonged incubation in a reaction with cellopentaose, and soluble cellooligosaccharides, such as cellobiose, cellotriose, cellotetraose, and cellohexaose, were also produced. No glucose was produced in the reaction even when it was carried out for a long time. The total number of molecules (cellooligosaccharides) in the reaction mixture remained at the initial substrate level during the entire reaction. The beta-transglycosylase proved to be a specific transferase showing transfer activity of glucosyl, cellobiosyl, and cellotriosyl moieties from one cellopentaose to an acceptor molecule from cellopentaose upwards with almost 100% efficiency.

Synthesis of levan by levansucrase. Some factors affecting the rate of synthesis and degree of polymerization of levan.

The addition of levan as a so-called "acceptor" accelerated the rate of polymerization of levan catalyzed by levansucrase [EC 2.4.1.10]. However, this effect was seen only under conditions of low ionic strength. Under conditions of high ionic strength, only levan with an average degree of polymerization (DP) of 120 was synthesized with a reasonable yield. Incorporation of [14C]fructose residues into levan of high molecular weight (DP 1,200) added as an "acceptor" at high ionic strength was slight. It is suggested that the DP of levan synthesized is generally regulated by ionic strength, and that enzymic synthesis of levan occurs in the absence of an "acceptor."

The molecular structure of low and high molecular weight levans synthesized by levansucrase.

The levan synthesized by Bacillus subtilis levansucrase in the presence of alcohols was of only high molecular weight, while in solutions of high ionic strength only low molecular weight (MW) levan was produced. The addition of low MW levan to the enzyme reaction mixture at low ionic strength stimulated synthesis of a high MW levan, but the levan added was not incorporated into this high MW levan. Methylation analysis revealed that low MW levans contained glucose, which was isolated as 2,3,46-tetra-O-methyl alditol acetate showing that the glucose units existed as terminal residues. The molecular weight of levan estimated on the basis of glucose content coincided with that determined by the gel filtration method. Methylation analysis also revealed that the number of fructose residues of the linear fraction linked by leads to 6(F)2 leads to type bonds was 22 for levan with a molecular weight of (8.4(-22)) x 10(3), while it was 11 for that of 2,000 x 10(3). The number of (formula: see text) type branched residues increased with increase in the molecular weight of the levan synthesized.

Potassium-stimulating mechanism of geranylgeranyl diphosphate synthase of Methanobacterium thermoformicicum SF-4.

The catalytic properties of geranylgeranyl diphosphate (GGPP) synthase [EC 2.5.1.29] purified from Methanobacterium thermoformicicum SF-4 were studied by kinetic procedures. The plots of 1/v versus 1/[S] and inhibition patterns by enzyme reaction products, PPi and GGPP, showed that the GGPP synthase reaction mechanism is an ordered-sequential Bi Bi one. Monovalent cations at low concentration (0.05 M) enhanced the enzyme activity, but at high concentration (0.4 M) they were inhibitory, except for K+. The K+ ion was found to be a modifier forming a parallel reaction pathway and accelerated the binding of substrates to the enzyme, especially the binding of isopentenyl diphosphate (IPP). When substrate concentrations are near the Km values, the rate-limiting step of the GGPP synthase reaction may be the substrate-binding step, probably the IPP-binding step, rather than the conversion step of the enzyme-farnesyl diphosphate-IPP complex to the enzyme-PPi-GGPP complex.

Structures of heterooligosaccharides synthesized by levansucrase.

In the presence of excess amounts of various monosaccharides as acceptors, Bacillus subtilis levansucrase synthesized various heterooligosaccharides as a result of transfer of the fructosyl residue from sucrose. The saccharides produced in the presence of D-glucose, D-mannose, and D-xylose were all non-reducing disaccharides and were identified as beta-D-fructofuranosyl-alpha-D-glucopyranoside (sucrose), beta-D-fructo-furanosyl-alpha-D-mannopyranoside (mannosucrose), and beta-D-fructofuranosyl-alpha-D-xylopyranoside (xylsucrose), respectively. The saccharides produced in the presence of D-galactose were also non-reducing, but consisted of di-, tri-, and tetrasaccharides. The di- and trisaccharides were galsucrose and 6F-beta-D-fructofuranosyl-galsucrose. The saccharide synthesized in the presence of L-arabinose was a reducing saccharide and was determined to be 4-O-beta-D-fructofuranosyl-L-arabinose. In the presence of D-fructose, several reducing levan oligomers, levanbiose, levantriose, levantetraose, etc., were produced, which were not observed in the synthesis of levan from sucrose alone.

Physiological activity of warburganal and its reactivity with sulfhydryl groups.

Warburganal, a unique dialdehyde sesquiterpene isolated from East African Warburgia plants, showed a strong antifungal activity. However, this growth inhibition in Saccharomyces cerevisiae was reversed with L-cysteine. In addition, warburganal inhibited the alcoholic fermentation of S. cerevisiae while L-cysteine reversed this inhibition. When alcohol dehydrogenase, a sulfhydryl enzyme, was incubated with warburganal, the enzyme activity decreased with time. The decrease was more rapid at alkaline pH. L-Cysteine prevented this enzyme inhibition by warburganal but could not restore the enzyme activity lost already due to warburganal. Warburganal lost its characteristic ultraviolet absorption spectrum in the presence of L-cysteine. The change in absorbance was favored at alkaline pH, indicating Michael reaction type addition of L-cysteine to warburganal. Based on these observations, a variety of physiological activities due to warburganal appear to result from its irreversible reactivity with sulfhydryl groups.

Aspartate aminotransferase from a thermophilic formate-utilizing methanogen, Methanobacterium thermoformicicum strain SF-4: relation to serine and phosphoserine aminotransferases, but not to the aspartate aminotransferase family.

The primary structure of the aspartate aminotransferase (AspAT) of an archaebacterium, Methanobacterium thermoformicicum strain SF-4, has been determined by cloning and sequencing of the gene for the enzyme. The gene had a consensus promoter and a ribosome binding sequence of methanogens in the 5' untranslated region, followed by an open reading frame starting with ATG and terminating with TGA. The deduced amino acid sequence was identical with the partial amino acid sequences of the enzyme including the N-terminal sequence, and the deduced molecular weight of 41,684 was virtually identical to that reported earlier for this enzyme [Tanaka, T., Yamamoto, S., Taniguchi, M., Hayashi, H., Kuramitsu, S., Kagamiyama, H., & Oi, S. (1992) J. Biochem. 112, 811-815]. The gene was expressed in Escherichia coli by inserting it into an expression vector just downstream of the lacZ promoter, and this verified that the cloned gene really encodes the Methanobacterium AspAT. The primary structure of the Methanobacterium AspAT showed extremely low homology, 5%, with AspATs of eubacteria, eukaryotes, and a thermoacidophilic arachaebacterium, Sulfolobus solfataricus. On the other hand, the Methanobacterium AspAT showed remarkable amino acid sequence homology, 31.5%, with rat serine:pyruvate aminotransferase and, 13.5%, with E. coli phosphoserine aminotransferase. Thus, the Methanobacterium AspAT apparently belongs to subgroup IV of the aminotransferases [Mehta, P.K., Hale, T.I., & Christen, P. (1993) Eur. J. Biochem. 214, 549-561], but not to subgroup I, in which all the AspATs known so far are included.

Further studies on aspartate aminotransferase of thermophilic methanogens by analysis of general properties, bound cofactors, and subunit structures.

Aspartate aminotransferase (AspAT) [EC 2.6.1.1] of thermophilic methanogen was further characterized with the enzyme from Methanobacterium thermoautotrophicum strain FTF-INRA as well as M. thermoformicicum strain SF-4. AspAT of strain FTF-INRA was similar in the amino donor specificity to the enzyme of M. thermoformicicum strain SF-4, in that it was active on L-cysteine and L-cysteine sulfinate in addition to L-glutamate and L-aspartate. The enzymes gave similar absorption spectra having maxima at around 326 and 415 nm with no pH-dependent shift but were found to contain 1 mol of tightly bound pyridoxal 5'-phosphate (PLP) per subunit. Reconstitution of each apoenzyme with added PLP resulted in partial recovery of the original enzymatic activity, suggesting a significant conformational change of the active site region upon removal of the cofactor. Polyacrylamide gel electrophoresis (PAGE) and gel filtration analyses revealed a tetrameric structure (180 kDa) of identical subunits with a molecular mass of 43 kDa for each of these enzymes. Electric current was found to affect the interaction or affinity of each subunit, promoting dissociation of the native enzyme into the monomeric form. Alkaline treatment was effective only for dissociation of the enzyme from strain SF-4. They were distinguishable by the more rapid reassociation of the monomer to the native aggregated form in the enzyme of strain FTF-INRA.

Ruthenium complex-catalyzed direct ortho arylation and alkenylation of 2-arylpyridines with organic halides.

[reaction: see text] The ortho position of the aromatic ring of pyridyl group-substituted aromatic compounds is directly arylated or alkenylated with organic halides in the presence of a catalytic amount of a ruthenium(II)-phosphine complex.

Lovastatin prevents angiotensin II-induced cardiac hypertrophy in cultured neonatal rat heart cells.

Angiotensin II activates p21ras, and mediates cardiac hypertrophic growth through the type 1 angiotensin II receptor in cardiac myocytes. An inhibitor of 3-hydroxy-3-methyglutaryl-coenzyme A (HMG-CoA) reductase has been shown to block the post-translational farnesylation of p21ras and inhibit protein synthesis in several cell types. Primary cultures of neonatal cardiac myocytes were used to determine whether HMG-CoA reductase inhibitors, lovastatin, simvastatin and pravastatin inhibit the angiotensin II-induced hypertrophic growth. Angiotensin II (10(-6) M) significantly increased protein-DNA ratio, RNA-DNA ratio, ratios of protein synthesis and mitogen-activated protein (MAP) kinase activity. Lipid-soluble HMG-CoA reductase inhibitors, lovastatin (10(-6) M) and simvastatin (10(-6) M) partially and significantly inhibited the angiotensin II-induced increases in these parameters, but a water-soluble HMG-CoA reductase inhibitor, pravastatin (10(-6) M) did not. Mevalonate (10(-4) M) overcame the inhibitory effects of lovastatin and simvastatin on angiotensin II-induced increases in these parameters. A selective protein kinase C inhibitor, calphostin C (10(-6) M) partially and significantly prevented angiotensin II-induced increases in these parameters, and treatment with both lovastatin and calphostin C inhibited completely. Angiotensin II increased p21ras activity and membrane association, and lovastatin inhibited them. These studies demonstrate that a lipid-soluble HMG-CoA reductase inhibitor, lovastatin, may prevent angiotensin II-induced cardiac hypertrophy, at least in part, through p21ras/MAP kinase pathway, which is linked to mevalonate metabolism.

Pathophysiology of nonneoplastic obstruction of the foramen of Monro and progressive unilateral hydrocephalus.

Our experience with progressive unilateral hydrocephalus in children and the pathophysiology of nonneoplastic obstruction of the foramen of Monro has been analyzed. The status of the foramen of Monro in either congenital or acquired progressive unilateral hydrocephalus was classified into the following four categories: Category 1, atresia of the foramen; Category 2, morphological obstruction; Category 3, functional obstruction; and Category 4, patent foramen. Illustrative cases including hydrodynamic studies and intracranial pressure monitoring are discussed.

Spontaneous reduction of a recurrent craniopharyngioma in an 8-year-old female patient: case report.

OBJECTIVE AND IMPORTANCE: The spontaneous rupture of a craniopharyngioma is an extremely rare condition confined to adults. This is the first report of a patient younger than 10 years who experienced spontaneous reduction (possibly rupture) of a craniopharyngioma. CLINICAL PRESENTATION: An 8-year-old female patient with a recurrence of a craniopharyngioma experienced fever, headache, and visual disturbance that lasted a few days. Concurrent with the improvement of these symptoms, marked reduction in the size of the tumor was revealed using magnetic resonance imaging, suggesting the occurrence of a rupture. INTERVENTION: Subsequent magnetic resonance imaging of the hypothalamic-pituitary region was performed while the patient received growth hormone therapy. CONCLUSION: There was no increase in the size of the tumor 1 year after the reduction occurred. Prompt evaluation of the hypothalamic-pituitary region using magnetic resonance imaging is warranted to rule out the possibility of spontaneous reduction (including rupture) of the tumor in a situation in which the patient with a craniopharyngioma shows meningeal signs or a rapid change of neurological symptoms (such as headache, fever, or visual disturbance).