Search PubMed⌕ Search

Biomedical subjects

O Adachi

Publications and source records attributed to O Adachi.

At least 73 records · Page 4Linked to original sources
PubMed1991

Reconstitution of the ethanol oxidase respiratory chain in membranes of quinoprotein alcohol dehydrogenase-deficient Gluconobacter suboxydans subsp. alpha strains.

The ethanol oxidase respiratory chain of Gluconobacter suboxydan was characterized by using G. suboxydans subsp. alpha, a variant species of G. suboxydans incapable of oxidizing ethanol. The membranes of G. suboxydans subsp. alpha exhibited neither alcohol dehydrogenase, ethanol oxidase, nor glucose-ferricyanide oxidoreductase activity. Furthermore, the respiratory chain of the organism exhibited an extremely diminished amount of cytochrome c and an increased sensitivity of the respiratory activity for cyanide or azide when compared with G. suboxydans. The first-subunit quinohemoprotein and the second-subunit cytochrome c of alcohol dehydrogenase complex in the membranes of G. suboxydans subsp. alpha were shown to be reduced and deficient, respectively, by using heme-staining and immunoblotting methods. Ethanol oxidase activity, lacking in G. suboxydans subsp. alpha, was entirely restored by reconstituting alcohol dehydrogenase purified from G. suboxydans to the membranes of G. suboxydans subsp. alpha; this also led to restoration of the cyanide or azide insensitivity and the glucose-ferricyanide oxidoreductase activity in the respiratory chain without affecting other respiratory activities such as glucose and sorbitol oxidases. Ethanol oxidase activity was also reconstituted with only the second-subunit cytochrome c of the enzyme complex. The results indicate that the second-subunit cytochrome c of the alcohol dehydrogenase complex is essential in ethanol oxidase respiratory chain and may be involved in the cyanide- or azide-insensitive respiratory chain bypass of G. suboxydans.

Acetobacter↗
PubMed1990

Cytochrome a1 of acetobacter aceti is a cytochrome ba functioning as ubiquinol oxidase.

Cytochrome a1 is a classic cytochrome that in the 1930s had already been detected in Acetobacter strains and in the 1950s was identified as a terminal oxidase. However, recent studies did not substantiate the previous observations. We have detected a cytochrome a1-like chromophore in Acetobacter aceti, which was purified and characterized in this study. The cytochrome was solubilized from membranes of the strain with octyl beta-D-glucopyranoside and was purified by single column chromatography. The purified cytochrome exhibited a broad alpha peak around 600-610 nm, which turned to a sharp peak at 589 nm in the presence of cyanide. Carbon monoxide difference spectra of the cytochrome indicated the presence of an alpha-type cytochrome. The cytochrome contained 1 mol each of hemes b and a and probably one copper ion. These results suggest that the cytochrome purified from A. aceti is the so-called cytochrome a1, and thus the existence of the classic cytochrome has been reconfirmed. The purified enzyme consisted of four polypeptides of 55, 35, 22, and 18 kDa, and it showed a sedimentation coefficient of 6.3 S in the native form. The enzyme had a high ubiquinol oxidase activity (140-160 mumol of ubiquinol-2 oxidized per min per mg of protein). When reconstituted into proteoliposomes, the cytochrome could generate an electrochemical proton gradient during oxidation of ubiquinol. Thus, cytochrome a1 of A. aceti has been shown to be a cytochrome ba terminal oxidase capable of generating an electrochemical proton gradient concomitant with ubiquinol oxidation.

Acetobacter↗
PubMed1990

Evidence for electron transfer via ubiquinone between quinoproteins D-glucose dehydrogenase and alcohol dehydrogenase of Gluconobacter suboxydans.

Gluconobacter suboxydans contains membrane-bound D-glucose and alcohol dehydrogenases (GDH and ADH) as the primary dehydrogenases in the respiratory chain. These enzymes are known to be quinoproteins having pyrroloquinoline quinone as the prosthetic group. GDH reduces an artificial electron acceptor, ferricyanide, in the membrane, but not after solubilization with Triton X-100, while ADH can react with the electron acceptor even after solubilization and further purification. In this study, it has been shown that the ferricyanide reductase activity of GDH is restored by adding the supernatant solubilized with Triton X-100 to the residue, and also by incorporation of purified ADH into the membranes of an ADH-deficient strain. G. suboxydans var. alpha. In addition, the ferricyanide reductase activity of GDH was reconstituted in proteoliposomes from GDH, ADH, and ubiquinone-10. Thus, the results indicated that the electron transfer from GDH to ferricyanide was mediated by ubiquinone and ADH. The data also suggest that GDH and ADH transfer electrons mutually via ubiquinone in the respiratory chain.

Alcohol Dehydrogenase↗
PubMed1990

New biological properties of pyrroloquinoline quinone and its related compounds: inhibition of chemiluminescence, lipid peroxidation and rat paw edema.

Pyrroloquinoline quinone (PQQ) inhibited the chemiluminescence (CL) from mouse peritoneal cells initiated by zymosan, carrageenin and N-formyl-methionyl-leucyl-phenylalanine and CL generated by the xanthine-xanthine oxidase reaction and the lipid peroxidation in the rat brain homogenate. The inhibitory activity of PQQ was more potent than that of idebenone, alpha-tocopherol and ascorbic acid in all the three assay systems. In the xanthine-xanthine oxidase reaction, PQQ had no effect on the formation of uric acid at the concentration of CL inhibition. These results suggest that PQQ might have a radical scavenger-like activity. Structure-activity relationship of PQQ and its six related compounds showed that the 7- and 9-carboxyl groups of PQQ as well as the orthoquinone structure are responsible for the radical scavenger-like activity. In addition, the -NH group in the pyrrole ring of PQQ seemed to be essential for the antilipid peroxidative activity in the rat brain homogenate. When administered i.p., PQQ inhibited the development of 0.1% carrageenin-induced paw edema in rats. These results suggest that PQQ might have therapeutic effects on various diseases, of which development or exacerbation has been known to be associated with radical oxygens.

Animals↗
PubMed1989

Quinoprotein D-glucose dehydrogenase of the Acinetobacter calcoaceticus respiratory chain: membrane-bound and soluble forms are different molecular species.

Acinetobacter calcoaceticus is known to contain soluble and membrane-bound quinoprotein D-glucose dehydrogenases, while other oxidative bacteria contain the membrane-bound enzyme exclusively. The two forms of glucose dehydrogenase were believed to be the same enzyme or interconvertible forms. Previously, Matsushita et al. [(1988) FEMS Microbiol. Lett 55, 53-58] showed that the two enzymes are different with respect to enzymatic and immunological properties, as well as molecular weight. In the present study, we purified both enzymes and compared their kinetics, reactivity with ubiquinone homologues, and immunological properties in detail. The purified membrane-bound enzyme had a molecular weight of 83,000, while the soluble form was 55,000. The purified enzymes exhibited totally different enzymatic properties, particularly with respect to reactivity toward ubiquinone homologues. The soluble enzyme reacted with short-chain homologues only, whereas the membrane-bound enzyme reacted with long-chain homologues including ubiquinone 9, the native ubiquinone of the A. calcoaceticus. Furthermore, the two enzymes were distinguished immunochemically; the membrane-bound enzyme did not cross-react with antibody raised against the soluble enzyme, nor did the soluble enzyme cross-react with antibody against the membrane-bound enzyme. Thus, each glucose dehydrogenase is a molecularly distinct entity, and the membrane-bound enzyme only is coupled to the respiratory chain via ubiquinone.

Acinetobacter↗
PubMed1989

Quinoprotein D-glucose dehydrogenases in Acinetobacter calcoaceticus LMD 79.41: purification and characterization of the membrane-bound enzyme distinct from the soluble enzyme.

Acinetobacter calcoaceticus is known to contain soluble and membrane-bound quinoprotein D-glucose dehydrogenases while other oxidative bacteria such as Pseudomonas or Gluconobacter contain only membrane-bound enzyme. The two different forms were believed to be the same enzyme or interconvertible. Present results show that the two different forms of glucose dehydrogenase are distinct from each other in their enzymatic and immunological properties as well as in their molecular size. The soluble and membrane-bound glucose dehydrogenases were separated after French press-disruption by repeated ultracentrifugation, and then purified to nearly homogeneous state. The soluble enzyme was a polypeptide of 55 Kdaltons, while the membrane-bound enzyme was a polypeptide of 83 Kdaltons which is mainly monomeric in detergent solution. Both enzymes showed different enzymatic properties including substrate specificity, optimum pH, kinetics for glucose, and reactivity for ubiquinone-homologues. Furthermore, the two enzymes could be distinguished immunochemically; the membrane-bound enzyme is cross-reactive with an antibody raised against membrane-bound enzyme purified from Pseudomonas but not with antibody elicited against the soluble enzyme, while the soluble enzyme is not cross-reactive with the antibody of membrane-bound enzyme. Data also suggest that the membrane-bound enzyme functions by linking to the respiratory chain via ubiquinone though the function of the soluble enzyme remains unclear.

Acinetobacter↗
PubMed1989

Reactivity with ubiquinone of quinoprotein D-glucose dehydrogenase from Gluconobacter suboxydans.

D-Glucose dehydrogenase is a pyrroloquinoline quinone-dependent oxidoreductase linked to the respiratory chain of a wide variety of bacteria. There is a controversy as to whether the glucose dehydrogenase is linked to the respiratory chain via ubiquinone or cytochrome b. In this study, it was shown that the glucose dehydrogenase of Gluconobacter suboxydans has the ability to react directly with ubiquinone. The enzyme purified from the membranes of G. suboxydans was able to react with ubiquinone homologues such as ubiquinone-1, -2, or -6 in detergent solution. Furthermore, in order to demonstrate the reactivity of the enzyme with native ubiquinone, ubiquinone-10, in the native membranous environment, the dehydrogenase was reconstituted together with cytochrome o, the terminal oxidase of the respiratory chain, into a phospholipid bilayer containing ubiquinone-10. The proteoliposomes thus reconstituted exhibited a reasonable glucose oxidase activity, the electron transfer reaction of which was able to generate a membrane potential and a pH gradient. Thus, D-glucose dehydrogenase of G. suboxydans has been demonstrated to donate electrons directly to ubiquinone in the respiratory chain.

Carbohydrate Dehydrogenases↗
PubMed1988

Pyrroloquinoline quinone: excretion by methylotrophs and growth stimulation for microorganisms.

A marked excretion of pyrroloquinoline quinone (PQQ) by methylotrophs into the culture medium was observed when incubation was prolonged to the late stationary phase. When the organisms were growing vigorously in the early exponential phase, accumulation of PQQ was repressed at a low level. Some evidence was obtained that the excretion of PQQ is related to turnover of quinoproteins of the organisms. The growth stimulation of microorganisms by PQQ was demonstrated using Acetobacter aceti. The presence of PQQ even at the pg/ml level in the culture medium stimulated the bacterial growth by reducing the lag time. The growth stimulating effect of PQQ was observed only by the reduction of the lag time but not by increase in either the subsequent growth rate or the total cell yield. The results indicated that PQQ must have an important role in the initiation of cell reproduction.

Acetobacter↗
PubMed1988

Adduct formation of pyrroloquinoline quinone and amino acid.

When pyrroloquinoline quinone (PQQ) is mixed with an amino acid, a corresponding Schiff base PQQ adduct is readily formed between carbonyl groups of PQQ and the primary amino group. A potent growth stimulating effect for microorganisms was observed with the PQQ adduct when it was administered in a culture medium. Although PQQ itself shows a marked growth stimulating effect, PQQ adducts appeared to be more active than authentic PQQ when compared on a molar basis. Conversely, unlike authentic PQQ, PQQ adducts were shown to be less active (greater than or equal to 100-fold) as the prosthetic group for a quinoprotein apo-glucose dehydrogenase when examined by holoenzyme formation by exogenous addition of PQQ or PQQ adducts. These observations suggested that PQQ adduct formation readily occurs during isolation procedures for PQQ from biological materials or PQQ - chromophore from quinoproteins. Therefore, the presence of such adducts gives a PQQ estimation much lower than theoretically expected. As an example, formation, isolation and characterization of PQQ - serine are described.

Amino Acids↗
PubMed1987

Reconstitution of pyrroloquinoline quinone-dependent D-glucose oxidase respiratory chain of Escherichia coli with cytochrome o oxidase.

D-Glucose dehydrogenase is a pyrroloquinoline quinone-dependent primary dehydrogenase linked to the respiratory chain of a wide variety of bacteria. The enzyme exists in the membranes of Escherichia coli, mainly as an apoenzyme which can be activated by the addition of pyrroloquinoline quinone and magnesium. Thus, membrane vesicles of E. coli can oxidize D-glucose to gluconate and generate an electrochemical proton gradient in the presence of pyrroloquinoline quinone. The D-glucose oxidase-respiratory chain was reconstituted into proteoliposomes, which consisted of two proteins purified from E. coli membranes, D-glucose dehydrogenase and cytochrome o oxidase, and E. coli phospholipids containing ubiquinone 8. The electron transfer rate during D-glucose oxidation and the membrane potential generation in the reconstituted proteoliposomes were almost the same as those observed in the membrane vesicles when pyrroloquinoline quinone was added. The results demonstrate that the quinoprotein, D-glucose dehydrogenase, can reduce ubiquinone 8 directly within phospholipid bilayer and that the D-glucose oxidase system of E. coli has a relatively simple respiratory chain consisting of primary dehydrogenase, ubiquinone 8, and a terminal oxidase.

Electron Transport Complex IV↗
PubMed1986

The 9-carboxyl group of pyrroloquinoline quinone, a novel prosthetic group, is essential in the formation of holoenzyme of D-glucose dehydrogenase.

Availability of different analogues of pyrroloquinoline quinone as the prosthetic group for apo-D-glucose dehydrogenase was examined. The 9-carboxyl group of pyrroloquinoline quinone was shown to be essential for the reconstitution of the enzyme activity. Although the carboxyl group may not be involved in catalytic function, it is quite probable to contribute the binding of the prosthetic group to apoenzyme.

Carbohydrate Dehydrogenases↗
PubMed1985

Identification of the covalently bound flavins of D-gluconate dehydrogenases from Pseudomonas aeruginosa and Pseudomonas fluorescens and of 2-keto-D-gluconate dehydrogenase from Gluconobacter melanogenus.

An improved method is presented for the purification of 8 alpha-(N1-histidyl)riboflavin, 8 alpha-(N3-histidyl)riboflavin and their 2',5'-anhydro forms, which permits the isolation of sizeable quantities of each of these compounds from a synthetic mixture in pure form. Flavin peptides were isolated from the D-gluconate dehydrogenases of Pseudomonas aeruginosa and Pseudomonas fluorescens and from the 2-keto-D-gluconate dehydrogenase of Gluconobacter melanogenus. After conversion into the aminoacyl-riboflavin, the flavin in all three enzymes was identified as 8 alpha-(N3-histidyl)riboflavin. By sequential treatment with nucleotide pyrophosphatase and alkaline phosphatase, the flavin in each enzyme was shown to be in the dinucleotide form.

Carbohydrate Dehydrogenases↗
PubMed1985

Method of enzymatic determination of pyrroloquinoline quinone.

An improved enzymatic method for the determination of pyrroloquinoline quinone, a novel prosthetic group of some important oxidoreductases, has been developed with cytoplasmic membrane of Escherichia coli K-12, in which D-glucose dehydrogenase (EC 1.1.99.17) was completely resolved to apo-enzyme by EDTA treatment. Incubation of the EDTA-treated membrane with exogenous pyrroloquinoline quinone in the presence of magnesium ions gave a quantitative determination of pyrroloquinoline quinone by assaying the restored D-glucose dehydrogenase activity. This novel enzymatic method was confirmed to be highly reproducible up to 10 ng of pyrroloquinoline quinone and could be applied to a routine assay of pyrroloquinoline quinone.

Cell Membrane↗
PubMed1985

Enzymatic determination of serum ethanol with membrane-bound dehydrogenase.

In this enzymatic method for detecting ethanol in blood by use of membrane-bound microbial alcohol dehydrogenase (no EC no. assigned), the enzyme catalyzes the reaction irreversibly and the rate of oxidation can be monitored by spectrophotometry of the reduction of the indicator dye. No pyridine nucleotides such as NAD+ or NADP+ are used. The calibration curve is linear in the range of 0.1 to 4.0 g of ethanol per liter. Assays of 45 samples of serum having ethanol values ranging from 0.4 to 3.2 g/L by the described technique and a gas-chromatographic method gave respective means of 1.734 and 1.732 g/L (r = 0.954).

Alcohol Dehydrogenase↗
PubMed1983

Membrane-bound respiratory chain of Pseudomonas aeruginosa grown aerobically. A KCN-insensitive alternate oxidase chain and its energetics.

There was found to be a KCN-insensitive, alternate oxidase chain branching from the ordinary oxidase chain in the respiratory chain of Pseudomonas aeruginosa grown aerobically. The alternate oxidase activity was highly resistant to KCN, and had a lower affinity for oxygen than ordinary cytochrome oxidase did. The branching point of the alternate oxidase chain from the ordinary oxidase chain was shown to be localized behind cytochrome b. The KCN-insensitive alternate oxidase chain was inhibited slightly with antimycin A and intensively with 2-thenoyltrifluoroacetone. The former inhibited the respiration behind cytochrome b and the latter before cytochrome b. N,N,N',N'-Tetramethyl-p-phenylenediamine oxidase-negative mutant (T105) was prepared from P. aeruginosa. The mutant clearly lacked a functional ordinary cytochrome oxidase, but had the KCN-insensitive alternate oxidase chain and could grow aerobically. The KCN-insensitive alternate oxidase chain had a H+/O ratio of 4, suggesting the existence of two energy-coupling sites in the chain. Under the conditions where both ordinary oxidase and alternate oxidase chains were functioning, the H+/O ratio of the parent strain was 5.6. From these data, we also discuss the energetics of the ordinary oxidase chain in the respiratory chain of aerobic P. aeruginosa.

Aerobiosis↗