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O Adachi

Publications and source records attributed to O Adachi.

87 records · Page 5Linked to original sources
PubMed1982

Membrane-bound cytochromes c of Pseudomonas aeruginosa grown aerobically. Purification and characterization of cytochromes c-551 and c-555.

Membrane-bound cytochromes c of Pseudomonas aeruginosa grown aerobically were investigated. By detecting polypeptides with heme-catalyzing peroxidase activity on a sodium dodecyl sulfate polyacrylamide gel, four major (Band I, 33,000 daltons; II, 25,000; III, 20,000; IV, 16,000) and one minor (V, 11,500) hemoproteins were found in the membrane fraction, while one hemoprotein (VI, 8,200) was detected in a small amount in the cytosol fraction. All these hemoproteins (bands I to VI) appeared to be cytochromes c, because all bands were detected even after being treated with HCl-acetone. Of the membrane-bound cytochromes c, cytochromes c-551 (band IV) and c-555 (band V) were solubilized with Triton X-100 and purified by repeated DEAE-cellulose column chromatography. Both purified cytochromes c-551 and c-555 were monomeric and their molecular weights were estimated to be 16,400 and 11,500, respectively. Their respective midpoint potentials were 0.31 and 0.34 V, and their respective isoelectric points in the reduced form were 3.8 and 5.2. The purified cytochromes c-551 and c-555 were found to be clearly different from "soluble" cytochrome c-551, and might function in the membrane-bound aerobic respiratory chain of P. aeruginosa.

Amino Acids↗
PubMed1981

D-fructose dehydrogenase of Gluconobacter industrius: purification, characterization, and application to enzymatic microdetermination of D-fructose.

D-Fructose dehydrogenase was solubilized and purified from the membrane fraction of glycerol-grown Gluconobacter industrius IFO 3260 by a procedure involving solubilization of the enzyme with Triton X-100 and subsequent fractionation on diethylaminoethyl-cellulose and hydroxylapatite columns. The purified enzyme was tightly bound to a c-type cytochrome and another peptide existing as a dehydrogenase-cytochrome complex. The purified enzyme was deemed pure by analytical ultracentrifugation as well as by gel filtration on a Sephadex G-200 column. The molecular weight of the enzyme complex was determined to be about 140,000, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed the presence of three components having molecular weights of 67,000 (dehydrogenase), 50,800 (cytochrome c), and 19,700 (unknown function). Only D-fructose was readily oxidized by the enzyme in the presence of dyes such as ferricyanide, 2,6-dichlorophenolindophenol, or phenazine methosulfate. Nicotinamide adenine dinucleotide, nicotinamide adenine dinucleotide phosphate, and oxygen did not function as electron acceptors. The optimum pH of D-fructose oxidation was 4.0. The enzyme was stable at pH 4.5 to 6.0 Stability of the purified enzyme was much enhanced by the presence of detergent in the enzyme solution. Removal of detergent from the enzyme solution facilitated the aggregation of the enzyme and caused its inactivation. An apparent Michaelis constant for D-fructose was observed to be 10(-2) M with the purified enzyme. D-Fructose dehydrogenase was shown to be a satisfactory reagent for microdetermination of D-fructose.

Carbohydrate Dehydrogenases↗
PubMed1980

Function of ubiquinone in the electron transport system of Pseudomonas aeruginosa grown aerobically.

The location and function of ubiquinone in the electron transport system of Pseudomonas aeruginosa grown aerobically were studied. The reduction level of ubiquinone in the intact membrane was 36-43% in the aerobic steady state and about 65% in the anaerobic state with one substrate, but the level in the anaerobic state reached to 81% with a mixture of several substrates. Complete removal of ubiquinone performed by extracting the lyophilized membrane particles with n-pentane containing acetone resulted in complete loss of all oxidase activities for glucose, gluconate, malate, succinate, and NADH. In the ubiquinone-depleted particles, neither cytochrome component was reduced by adding any substrate. Reincorporation of coenzyme Q9 into the depleted particles restored each oxidase activity to 60 to 80% of the original and reduction of cytochromes with substrates. The reduction kinetics of cytochromes and effect of inhibitors showed that coenzyme Q9 was incorporated at the original site in the electron transport system. Exogenous coenzyme Q2 increased gluconate and malate oxidase activities and decreased glucose oxidase activity, when French-pressed membrane vesicles but not spheroplasts were used. Oxidizing activity for reduced coenzyme Q2 was also detected in the pressed vesicles but not in the spheroplasts. The present results showed that ubiquinone was indispensable and located prior to cytochromes in the electron transport system. Furthermore, the homogeneity and sidedness of ubiquinone in the cytoplasmic membrane of the organism are also discussed.

Aerobiosis↗
PubMed1980

Membrane-bound respiratory chain of Pseudomonas aeruginosa grown aerobically.

The electron transport chain of the gram-negative bacterium Pseudomonas aeruginosa, grown aerobically, contained a number of primary dehydrogenases and respiratory components (soluble flavin, bound flavin, coenzyme Q9, heme b, heme c, and cytochrome o) in membrane particles of the organism. Cytochrome o, about 50% of the b-type cytochrome, seemed to function as a terminal oxidase in the respiratory chain. The electron transport chain of P. aeruginosa grown aerobically was suggested to be lined up in order of primary dehydrogenase, b, c1, c, o, and oxygen.

Aerobiosis↗
PubMed1979

Occurrence of old yellow enzyme in Gluconobacter suboxydans, and the cyclic regeneration of NADP.

Old yellow enzyme system has been found in the cytosol fraction of Gluconobacter suboxydans. This is the first time that the enzyme has been found in organisms other than yeast cells. Old yellow enzyme [EC 1.6.99.1], D-glucose-6-phosphate dehydrogenase [EC 1.1.1.49], and catalase were isolated and crystallized separately from the organism. The old yellow enzyme from G. suboxydans showed catalytic and physicochemical properties almost identical with those of the enzyme from yeast cells. NADPH was specifically oxidized by the old yellow enzyme and the reduced enzyme was spontaneously reoxidized by atmospheric oxygen. The old yellow enzyme from G. suboxydans also contained FMN as a prosthetic group, and two mol of FMN were found per mol of enzyme (molecular weight, 88,000 as determined by gel filtration). In the oxidation of D-glucose-6-phosphate to 6-phospho-D-gluconate, cyclic regeneration of NADP occurred smoothly in the presence of D-glucose-6-phosphate dehydrogenase and catalase, even when a limited amount of NADP or NADPH was present in the reaction mixture.

Catalase↗
PubMed1979

Membrane-bound D-gluconate dehydrogenase from Pseudomonas aeruginosa. Purification and structure of cytochrome-binding form.

A membrane-bound D-gluconate dehydrogenase [EC 1.1.99.3] was solubilized from membranes of Pseudomonas aeruginosa and purified to a homogeneous state with the aid of detergents. The solubilized enzyme was a monomer in the presence of at least 0.1% Triton X-100, having a molecular weight of 138,000 on polyacrylamide gel electrophoresis or 124,000--131,000 on sucrose density gradient centrifugation. In the absence of Triton X-100, the enzyme became dimeric, having a molecular weight of 240,000--260,000 on sucrose density gradient centrifugation. Removal of Triton X-100 caused a decrease in enzyme activity. Enzyme activity was stimulated by addition of phospholipid, particularly cardiolipin, in the presence of Triton X-100. The enzyme had a cytochrome c1, c-554(551), which might be a diheme cytochrome, and it also contained a covalently bound flavin but not ubiquinone. In the presence of sodium dodecyl sulfate, the enzyme was dissociated into three components with molecular weights of 66,000, 50,000, and 22,000. The components of 66,000 and 50,000 daltons corresponded to a flavoprotein and cytochrome c1, respectively, but that of 22,000 dalton remained unclear as to its function.

Carbohydrate Dehydrogenases↗
PubMed1978

Isolation and characterization of outer and inner membranes from Pseudomonas aeruginosa and effect of EDTA on the membranes.

The outer and inner cytoplasmic membranes of Pseudomonas aeruginosa were separated as small and large membranes, respectively, from the cell envelope of this organism treated with lysozyme in Tris-chloride buffer containing sucrose and MgCl2 by differential centrifugation. The small membrane fraction contained predominantly 2-keto-3-deoxyoctonate (KDO), and little cytochromes or oxidase activities. The small membrane was composed of only 9 polypeptides and showed homogeneous small vesicles electron-microscopically. On the other hand, the large membrane fraction had high cytochrome contents and oxidase activities, and little KDO. The large membrane was composed of a number of polypeptides and showed large fragments or vesicles electron-microscopically. These results indicate that the small and large membranes are the outer and inner cytoplasmic membranes of P. aeruginosa, respectively. The isolated outer membrane showed a symmetrical protein peak with a density of 1.23 on sucrose density gradient centrifugation and the isolated inner membrane showed an unusually high density, probably due to association with ribosomes and extrinsic or loosely bound proteins. EDTA lowered the density of both membranes and caused lethal damage to the outer membrane, causing disintegration with the release of lipopolysaccharide (LPS), proteins and phospholipid.

Carbohydrate Dehydrogenases↗
PubMedDate not supplied

Cellular localization and metabolic function of n-butylamine-induced amine oxidases in the fungus Aspergillus niger AKU 3302.

Using transmission electron microscopy, the amine oxidase activity in Aspergillus niger AKU 3302 was localized to the outer side of the cell wall but not inside the cell using the cerium perhydroxide deposition method. The presence of cerium in the deposit was confirmed by energy-dispersive microanalysis of X-rays. Interestingly, immunocytochemical localization using gold labeling with a specific antibody indicated the presence of amine oxidase protein inside the cell wall and not only on the outer surface. Besides labeling of the cell wall, a high level of labeling was also observed inside the cell in what seemed to be secretory vesicle structures. It is proposed that the highly active amine oxidase AO-I is located in the cell wall and serves primarily as a detoxifying agent, preventing amines from entering and damaging the cell. The amine oxidation exhibits an interesting spatial orientation involving a release of toxic hydrogen peroxide into the extracellular space. The inactive amine oxidase protein located inside the cell is most probably the amine oxidase AO-II, found in cell homogenates. It is also likely that the less active AO-II is an improperly folded precursor of AO-I, which acquired low-level activity after cell homogenization in the presence of Cu(II) and oxygen due to autooxidative formation of topaquinone.

Amine Oxidase (Copper-Containing)↗