Exogenous quinones inhibit photosynthetic electron transfer in Chloroflexus aurantiacus by specific quenching of the excited bacteriochlorophyll c antenna.

In the photosynthetic green filamentous bacterium Chloroflexus aurantiacus, excitation energy is transferred from a large bacteriochlorophyll (BChl) c antenna via smaller BChl a antennas to the reaction center. The effects of substituted 1,4-naphthoquinones on BChl c and BChl a fluorescence and on flash-induced cytochrome c oxidation were studied in whole cells under aerobic conditions. BChl c fluorescence in a cell suspension with 5.4 microM BChl c was quenched to 50% by addition of 0.6 microM shikonin ((R)-2-(1-hydroxy-4-methyl-3-pentenyl)-5,8-dihydroxy-1, 4-naphthoquinone), 0.9 microM 5-hydroxy-1,4-naphthoquinone, or 4 microM 2-acetyl-3-methyl-1,4-naphthoquinone. Between 25 and 100 times higher quinone concentrations were needed to quench BChl a fluorescence to a similar extent. These quinones also efficiently inhibited flash-induced cytochrome c oxidation when BChl c was excited, but not when BChl a was excited. The quenching of BChl c fluorescence induced by these quinones correlated with the inhibition of flash-induced cytochrome c oxidation. We concluded that the quinones inhibited electron transfer in the reaction center by specifically quenching the excitation energy in the BChl c antenna. Our results provide a model system for studying the redox-dependent antenna quenching in green sulfur bacteria because the antennas in these bacteria inherently exhibit a sensitivity to O(2) similar to the quinone-supplemented cells of Cfx. aurantiacus.     

Automated high-performance liquid chromatographic method for determination of furosemide in dog plasma

An automated high-performance liquid chromatographic method for the determination of the diuretic drug furosemide has been established. Dog plasma was injected directly into a two-column system with a BSA—ODS (ODS column coated with bovine serum albumin) precolumn and a C₁₈ analytical column for the separation of furosemide. The two columns were automatically switched. Furosemide remained trapped on the precolumn while proteins were eluted to waste. After column switching, furosemide was washed onto the analytical column and analysed without interference. The greatest advantage of the method is its easy performance without manual sample preparation; it requires no extraction or deproteinization. The method allows determination of 0.1–10 μg/ml of furosemide with accuracy and precision comparable with previously reported values. The coefficients of variation obtained from replicate measurements of 1 μg/ml and 5 μg/ml samples were 1.65% and 2.40%, respectively. This method was used to measure the plasma levels of furosemide in beagle dogs to whom the drugs was administered, as a reference, in a toxicological study.     

Nestmate recognition mediated by intestinal bacteria in a termite, Reticulitermes speratus

Discrimination and aggressive responses toward non-nestmates have been observed in a number of termite species, but the mechanism is poorly understood. Here I present the novel hypothesis that differential intestinal bacteria composition leads to production of colony-specific chemical cues that enable nestmate recognition.
The intestinal microflora of a lower termite, Reticulitermes speratus , consisted of many bacteria species. The composition of the intestinal bacteria was exclusively colony-specific. Termites that had adsorbed an unfamiliar odor of bacteria sampled from another colony were fiercely attacked by nestmates. Experimental manipulation of the composition of bacteria by antibiotics successfully changed the recognition behavior toward nestmates. These results indicate that intestinal bacteria play an important role in nestmate recognition.     

Simultaneous Determination of Oxidized and Reduced Forms of Plastoquinone A in Spinach Chloroplasts by High-Performance Liquid Chromatography with an Electrochemical Detector

A method for determination of the redox level of plastoquinoneA in spinach chloroplasts is described. Plastoquinone A andits reduced form plastoquinol A were extracted from chloroplastson a sample-preparation cartridge (SEP-PAK C₁₈ Cartridge, WatersAssoc. Inc.) with a mixture of ethanol and diethyl ether ( 1: 1, vv). Extracts were separated by reversed-phase high-performanceliquid chromatography and examined with an electrochemical detectorequipped with dual electrodes. Plastoquinone A was determinedby its reductive current on one electrode, and plastoquinolA by its oxidative current on the other electrode. This method was applied to the determination of the redox potentialof plastoquinone A in chloroplasts. The midpoint potential atpH 7.8 of plastoquinone A was +20 mV with an n number of 2. (Received March 30, 1987; Accepted August 3, 1987)     

Electron flow and heme-heme interaction between cytochromes b-558, b-595 and d in a terminal oxidase of Escherichia coli

The ESR signals of the cytochromes in the Escherichia coli terminal oxidase cytochrome d complex were studied at cryogenic temperature. The intensities and g values of the rhombic high-spin signals changed when the electronic state of cytochrome d was changed from the oxidized state to the reduced or oxygen-binding or CO-binding state. These rhombic signals were therefore assigned to cytochrome b-595, which is located near cytochrome d in the oxidase complex. This assignment was supported by the finding that the Em value of the rhombic signals differed from that of cytochrome d (Hata, A. et al. (1985) Biochim. Biophys. Acta 810, 62-72). Photolysis and ligand-exchange experiments with the reduced CO complex of the oxidase were performed in the presence of oxygen at -140 degrees C. The ESR spectra of three intermediate forms trapped by controlled low temperatures were detected. These forms were designated as the oxygen-binding intermediate I (ESR-silent), oxygen-binding intermediate II (giving ESR signals at g = 6.3, 5.5 and 2.15), and oxygen-binding intermediate III (giving signals at g = 6.3, 5.5 and 6.0). From these results, electron flow in the cytochrome d complex is proposed to proceed in the order, cytochrome b-558----cytochrome b-595----cytochrome d----O2. A model of the mechanism of four-electron chemistry for oxidation of ubiquinol-8 and formation of H2O by the cytochrome d complex is presented.     

Studies on DNA markers (D4S10 and D4S43/S127) genetically linked to Huntington's disease in Japanese families

Summary This is the first full report on the genetic linkage between Japanese Huntington's disease and the DNA markers D4S10 and D4S43/S127. With use of the HindIII, BglI, and EcoRI polymorphisms detected at D4S10, and the combination of all these polymorphisms to give composite haplotypes, nine Japanese Huntington's disease families were found to be informative. Three recombinants for D4S10 were detected in these families, giving a maximum lod score of 1.662 at a of 0.10. Similarly, when we used the MspI and PvuII polymorphisms detected by D4S43/S127, five families gave informative results. No recombinant was detected in these families, giving a maximum lod score of 3.348 at a of 0.00. These results clearly support the view that the Japanese Huntington's disease gene may be identical with the Western gene, in spite of the lower prevalence rate in Japan.     

Non-existence of a tight association between a 444leucine to proline mutation and phenotypes of Gaucher disease: high frequency of a NciI polymorphism in the non-neuronopathic form

Summary A ⁴⁴⁴leucine to proline mutation detected by a NciI polymorphism in the human glucocerebrosidase gene was studied to investigate the correlation of the three clinical phenotypes of Gaucher disease with this mutation in 11 Japanese patients with Gaucher disease (type I, 8 patients; type II, 1 patient; type III, 2 patients) and to determine the feasibility of the use of genomic probe DNA for carrier detection and prenatal diagnosis in 8 Japanese families with Gaucher disease and agreeable to family study (type I, 6 families; type III, 2 families). The homoallelic ⁴⁴⁴leucine to proline mutation was found only in patients with type I disease. Of the 8 type I patients, 5 had the homoallelic mutation and 2 had one mutant allele. One patient with type II disease did not have this mutant allele. Of the 2 type III patients, one had a single mutant allele whereas the other exhibited no mutation of this kind. These results suggest that the ⁴⁴⁴leucine to proline mutation is very common in the type I (non-neuronopathic form) disease and is not tightly associated only with neuronopathic types of Gaucher disease in Japanese patients. These findings seem to conflict with others showing that this mutation is partially responsible for the occurrence of neuronopathic Gaucher disease. Thus, the NciI polymorphism will not be useful for the diagnosis of subtypes of Gaucher disease. Carrier detection was feasible in three families with type I disease of the 8 families analyzed by the NciI polymorphism.     

Preferential ADP-ribosylation of arginine-3 in synthetic heptapeptide Leu-Arg-Arg-Ala-Ser-Leu-Gly.

Hen liver nuclear ADP-ribosyltransferase modified the synthetic heptapeptide Kemptide (Leu-Arg-Arg-Ala-Ser-Leu-Gly) at arginine-2 and/or arginine-3. Trypsin treatment of ADP-ribosyl-Kemptide revealed that the ADP-ribosylation of arginine-3 was constantly more abundant than that of arginine-2. ADP-ribosylation of Kemptide suppressed the subsequent phosphorylation by cyclic AMP-dependent protein kinase.     

Kinetic mechanism of pulmonary carbonyl reductase.

An IgG1 monoclonal antibody, Sulph I, reacting with sulphatide (3'-sulphogalactosylceramide), was produced by immunizing Balb/c mice with that glycolipid coated on Salmonella minnesota bacterial membrane. Radioimmunodetection of the binding of the monoclonal antibody to structurally related glycolipids adsorbed to microtitre plates or chromatographed on thin-layer plates was used to determine its binding epitope. The antibody showed similar binding avidity to three sulphated glycolipids: sulphatide, sulpholactosylceramide and seminolipid. Lysosulphatide did bind the antibody, but, compared with sulphatide, 30 times more antigen was needed for half-maximal binding. Bis(sulphogangliotriosyl)ceramide and bis-sulphogangliotetraosylceramide did not bind the antibody. These results suggest that terminal galactose-3-O-sulphate and part of the hydrophobic region of the glycolipid are recognized by the Sulph I antibody.