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3-Dehydroshikimate (3-DHS) is a key intermediate for the synthesis of various compounds, including the antiviral drug oseltamivir. The Gluconobacter oxydans strain NBRC3244 intrinsically oxidizes quinate to produce 3-dehydroquinate (3-DHQ) in the periplasmic space. Even though a considerable activity is detected in the recombinant G. oxydans homologously overexpressing type II dehydroquinate dehydratase (DHQase) encoded in the aroQ gene at a pH where it grows, an alkaline shift of the culture medium is required for 3-DHS production in the middle of cultivation. Here, we attempted to adopt type I DHQase encoded in the aroD gene of Gluconacetobacter diazotrophicus strain PAL5 because the type I DHQase works optimally at weak acid, which is preferable for growth conditions of G. oxydans. In addition, we anticipated that subcellular localization of DHQase is the cytoplasm, and therefore, transports of 3-DHQ and 3-DHS across the cytoplasmic membrane are rate-limiting steps in the biotransformation. The Sec- and TAT-dependent signal sequences for secretion were attached to the N terminus of AroD to change the subcellular localization. G. oxydans that expresses the TAT-AroD derivative achieved 3-DHS production at a tenfold higher rate than the reference strain that expresses wild-type AroD even devoid of alkaline shift. Enzyme activity with the intact cell suspension and signal sequence cleavage supported the relocation of AroD to the periplasmic space. The present study suggests that the relocation of DHQase improves 3-DHS production in G. oxydans and represents a proof of concept for the potential of enzyme relocation in metabolic engineering. Key points: • Type-I dehydroquinate dehydratase (DHQase) was expressed in Gluconobacter oxydans. • Cytoplasmic DHQase was relocated to the periplasmic space in G. oxydans. • Relocation of DHQase in G. oxydans improved 3-dehydroshikimate production. [ABSTRACT FROM AUTHOR]

Dihydroxyacetone (DHA), a chemical suntan agent, is produced by the regiospecific oxidation of glycerol with Gluconobacter thailandicus NBRC3255. However, this microorganism consumes DHA produced in the culture medium. Here, we attempted to understand the pathway for DHA metabolism in NBRC3255 to minimize DHA degradation. The two gene products, NBRC3255_2003 (DhaK) and NBRC3255_3084 (DerK), have been annotated as DHA kinases in the NBRC 3255 draft genome. Because the double deletion derivative for dhaK and derK showed ATP-dependent DHA kinase activity similar to that of the wild type, we attempted to purify DHA kinase from ∆dhaK ∆derK cells to identify the gene for DHA kinase. The identified gene was NBRC3255_0651, of which the product was annotated as glycerol kinase (GlpK). Mutant strains with several combinations of deletions for the dhaK, derK, and glpK genes were constructed. The single deletion strain ∆glpK showed approximately 10% of wild-type activity and grew slower on glycerol than the wild type. The double deletion strain ∆derK ∆glpK and the triple deletion strain ∆dhaK ∆derK ∆glpK showed DHA kinase activity less than a detection limit and did not grow on glycerol. In addition, although ΔderK ΔglpK consumed a small amount of DHA in the late phase of growth, ∆dhaK ΔderK ΔglpK did not show DHA consumption on glucose–glycerol medium. The transformants of the ∆dhaK ΔderK ΔglpK strain that expresses one of the genes from plasmids showed DHA kinase activity. We concluded that all three DHA kinases, DhaK, DerK, and GlpK, are involved in DHA metabolism of G. thailandicus. Key points: • Dihydroxyacetone (DHA) is produced but degraded by Gluconobacter thailandicus. • Phosphorylation rather than reduction is the first committed step in DHA metabolism. • Three kinases are involved in DHA metabolism with the different properties. [ABSTRACT FROM AUTHOR]

A novel oxidation of D-pentonates to 4-keto-D-pentonates was analyzed withGluconobacter thailandicusNBRC 3258. D-Pentonate 4-dehydrogenase activity in the membrane fraction was readily inactivated by EDTA and it was reactivated by the addition of PQQ and Ca2+. D-Pentonate 4-dehydrogenase was purified to two different subunits, 80 and 14 kDa. The absorption spectrum of the purified enzyme showed no typical absorbance over the visible regions. The enzyme oxidized D-pentonates to 4-keto-D-pentonates at the optimum pH of 4.0. In addition, the enzyme oxidized D-fructose to 5-keto-D-fructose, D-psicose to 5-keto-D-psicose, including the other polyols such as, glycerol, D-ribitol, D-arabitol, and D-sorbitol. Thus, D-pentonate 4-dehydrogenase was found to be identical with glycerol dehydrogenase (GLDH), a major polyol dehydrogenase inGluconobacterspecies. The reaction versatility of quinoprotein GLDH was notified in this study. [ABSTRACT FROM AUTHOR]

l-Phenylalanine ammonia-lyase (PAL, EC 4.3.1.25) from Rhodosporidium toruloides was utilized to remove l-phenylalanine ( l-Phe) from different commercial protein hydrolysates. A casein acid hydrolysate (CAH, l-Phe ~2.28 %) was employed as a model substrate. t-Cinnamic acid resulting from deamination of l-Phe was extracted, analyzed at λ = 290 nm, and used for PAL activity determination. Optimum reaction conditions, optimized using successive Doehlert design, were 35 mg mL of CAH and 800 mU mL of PAL, while temperature and pH were 42 °C and 8.7, respectively. Reaction kinetics of PAL with CAH was determined under optimized conditions. Then, removal of l-Phe from CAH was tested. Results showed that more than 92 % of initial l-Phe was eliminated. Similar results were obtained with other protein hydrolysates. These findings demonstrate that PAL is a useful biocatalyst for l-Phe removal from protein hydrolysates, which can be evaluated as potential ingredients in foodstuffs for PKU patients. [ABSTRACT FROM AUTHOR]

Shikimate and 3-dehydroshikimate are useful chemical intermediates for the synthesis of various compounds, including the antiviral drug oseltamivir. Here, we show an almost stoichiometric biotransformation of quinate to 3-dehydroshikimate by an engineered Gluconobacter oxydans strain. Even under pH control, 3-dehydroshikimate was barely detected during the growth of the wild-type G. oxydans strain NBRC3244 on the medium containing quinate, suggesting that the activity of 3-dehydroquinate dehydratase (DHQase) is the rate-limiting step. To identify the gene encoding G. oxydans DHQase, we overexpressed the gox0437 gene from the G. oxydans strain ATCC621H, which is homologous to the aroQ gene for type II DHQase, in Escherichia coli and detected high DHQase activity in cell-free extracts. We identified the aroQ gene in a draft genome sequence of G. oxydans NBRC3244 and constructed G. oxydans NBRC3244 strains harboring plasmids containing aroQ and different types of promoters. All recombinant G. oxydans strains produced a significant amount of 3-dehydroshikimate from quinate, and differences between promoters affected 3-dehydroshikimate production levels with little statistical significance. By using the recombinant NBRC3244 strain harboring aroQ driven by the lac promoter, a sequential pH adjustment for each step of the biotransformation was determined to be crucial because 3-dehydroshikimate production was enhanced. Under optimal conditions with a shift in pH, the strain could efficiently produce a nearly equimolar amount of 3-dehydroshikimate from quinate. In the present study, one of the important steps to convert quinate to shikimate by fermenting G. oxydans cells was investigated. [ABSTRACT FROM AUTHOR]

We succeeded in obtaining a strain adapted to higher temperature from a thermotolerant strain, Gluconobacter frateurii CHM43, for sorbose fermentation. The adapted strain showed higher growth and l-sorbose production than original CHM43 strain at higher temperature around 38.5-40 °C. It was also shown to be useful even with the fermentation without temperature control. To understand the sorbose fermentation ability of the adapted strain at higher temperature, d-sorbitol-oxidizing respiratory chain was compared with the CHM43 strain and the adapted strain. We found that the activity of pyrroloquinoline quinone (PQQ)-dependent glycerol dehydrogenase (GLDH), which is a primary dehydrogenase of the respiratory chain and responsible for l-sorbose production, was decreased when the temperature increased, but the decreased activity of GLDH was recovered by the addition of PQQ. Since the adapted strain was found to produce more PQQ than the CHM43 strain, it was suggested that the adapted strain keeps GLDH as holoenzyme with the increased PQQ production, and thus produces more l-sorbose and grows better under higher temperature. [ABSTRACT FROM AUTHOR]

The article discusses the findings of a study which determined the role of genes in oxidative fermentation of D-sorbitol in Gluconobacter frateurii bacteria. The researchers described the oxidizing characteristic of the Gluconobacter in producing L-sorbose, 2- and 5-keto-D-gluconates, and the Miglitol drug. The experiment confirmed the identification and characterization of D-sorbitol dehydrogenase genes including sldR, xdhA, and perA.

The article presents a study that attempts to identify the genes essential for growth at high temperature by transposon (Tn10) mutagenesis to elucidate the mechanism underlying the thermotolerance of mesophilic strains. It states that Acetobacter tropicalis SKU1100 is a thermotolerant acetic acid bacterium that dwells even at high temperatures. Results have further supported the partial overlap between the genes required for acetic acid resistance and those needed for thermotolerance.

The article presents a study concerning the formation of 4-Pentulosonates (4-Keto-D-pentonates) and 4-Keto-D-aldopentoses having unidentified membrane-bound enzymes coming from acetic acid bacteria. Study uses various methods to examine the formation including thin layer chromatography (TLC), centrifugation, and ethylenediaminetetraacetic acid (EDTA) treatment. Findings reveal the unidentified membrane-bound enzymes which contribute in the formation of 4-Pentulosonates and 4-Keto-D-aldopentoses.

The article presents a study on the production of 5-keto-D-gluconate (5KGA) derived from D-glucose by Gluconobacter. The study indicates that the effect of potential of hydrogen (pH) is essential in the production of 5KGA is correlative with the pH dependency of 5KGA. It notes that the 5KGA compound holds potential as an intermediate in the food industry.

The article focuses on the production of 4-keto-D-arabonate (4KAB) by Gluconacetobacter liquefaciens strains. It states that the strains accumulated 2,5-diketo-d-gluconate (25DKA) and oxidized D-gluconate, D-glucose, and 2-keto-D-gluconate (2KGA). It says that after the accumulation, 25DKA was converted into 4KAB and 4KAB remained stable in Gluconacetobacter liquefaciens. Moreover, a study of M. Ameyama and K. Kondo on carbohydrate metabolism of acetic acid bacteria assumed the 4KAB occurrence.

The article presents a study regarding the productiuon of 3-dehydroshikimate (DSA) from quinate by immobilixing the membrane fraction of the acetic acid bacteria Gluconobacter oxydans IFO 3244. The study used several methods to realize and examine the immobilization process including encapsulation, ionic binding, and physical adsorption. Results show a conversion rate of almost 100% from quinate to 3-dehydroshikimate.

The article presents a study concerning acetic acid fermentation of Acetobacter pasteurianus. The study involves the examination of the relationship between pellicle polysaccharide formation and acetic acid resistance. It shows that acetic acid resistance ability pellicle formation is primarily related to pellicle formation, thus is essential to acetic acid fermentation in A. pasteurianus strains.

The article presents a study that focuses on the purification and characterization of the membrane-bound 3-dehydroshikimate dehydratase (mDSD) and enzyme solubilization from the membrane fraction of gluconobacter oxydans IFO 3244. The results revealed that the mDSD in the membrane fraction suggests the mDSD's significance of aromatization, solubilized and purified to high homogeneity. It says that dehydroshikimate (DSA) was confirmed as a direct precursor of phenylcyclohexylamine (PCA).

We isolated several thermotolerant Acetobacter species of which MSU10 strain, identified as Acetobacter pasteurianus, could grow well on agar plates at 41°C, tolerate to 1.5% acetic acid or 4% ethanol at 39°C, similarly seen with A. pasteurianus SKU1108 previously isolated. The MSU10 strain showed higher acetic acid productivity in a medium containing 6% ethanol at 37°C than SKU1108 while SKU1108 strain could accumulate more acetic acid in a medium supplemented with 4–5% ethanol at the same temperature. The fermentation ability at 37°C of these thermotolerant strains was superior to that of mesophilic A. pasteurianus IFO3191 strain having weak growth and very delayed acetic acid production at 37°C even at 4% ethanol. Alcohol dehydrogenases (ADHs) were purified from MSU10, SKU1108, and IFO3191 strains, and their properties were compared related to the thermotolerance. ADH of the thermotolerant strains had a little higher optimal temperature and heat stability than that of mesophilic IFO3191. More critically, ADHs from MSU10 and SKU1108 strains exhibited a higher resistance to ethanol and acetic acid than IFO3191 enzyme at elevated temperature. Furthermore, in this study, the ADH genes were cloned, and the amino acid sequences of ADH subunit I, subunit II, and subunit III were compared. The difference in the amino acid residues could be seen, seemingly related to the thermotolerance, between MSU10 or SKU1108 ADH and IFO 3191 ADH. [ABSTRACT FROM AUTHOR]

The article discusses membrane-bound glucono-δ-lactonase (MGL) properties, solubility, and enzyme purification. It presents the process wherein MGL is purified from Gluconobacter oxydans. It also discusses that the involved acetic acid bacteria and aerobic bacteria in the oxidative fermentation contains a well-developed periplasmic oxydase system which yields bioenergy formation.

Chlorogenate hydrolase (EC 3.1.1.42, CHase) was highly induced in mycelia of Aspergillus sojae AKU 3312 grown in Czapek medium containing either instant coffee powder or coffee pulp as inducer. No CHase formation was observed in the mycelia when cultivated without the inducer. CHase was purified readily from CHase-induced mycelia to high homogeneity, and the purified CHase revealed the molecular weight of 180,000 consisting of two identical subunits of 88 kDa. Equimolar quinate (QA) and caffeate (CA) were confirmed on hydrolysis of chlorogenate (CGA). The purified CHase was only useful for a laboratory scale hydrolysis of CGA. For practical QA and CA production using scaled up hydrolysis of vegetable extracts of natural CGA resources, the enzyme activity of purified CHase decreased and denatured irreversibly. Preparation of coffee pulp koji and its application to QA and CA production were proposed instead of purified CHase. When coffee pulp koji was heated at 60°C for 30 min, CHase survived without any appreciable loss of enzyme activity while vegetative mycelial growth and spore germination were terminated. The heated coffee pulp koji thus prepared was effective itself as stable immobilized catalyst of CHase for QA and CA production from vegetable CGA resources such as coffee powders, coffee pulp, and others. [ABSTRACT FROM AUTHOR]

The article studies tightly-bound quinone functions in the ubiquinone reaction sites of the quinoprotein alcohol dehydrogenase (ADH) of the Gluconobacter suboxydans bacterium in acetic acid. It includes an in-depth analysis of the solubilization and purification of ADH with dodecyl maltoside, as well as a discussion on the issues' implications for biological chemistry.

The article presents a study which examines the purification and properties of dihydroxyacetone (DHA) reductases in gluconobacter suboxydans grown in gycerol. It states that two DHA reductases were detected in the same cytoplasm gluconobacter suboxydans. It notes that NADH-dependent appears inducible and labile in nature while the NADPH-dependent appears constitutive and stable. It also notes that the two DHARS were specific only to DHA reduction to glycerol.

The article reports on a study on the novel 3-dehydroquinate dehydrase (DQD) catalyzing extracellular formation of 3-dehydroshikimate by oxidative fermentation of Gluconobacter oxydans IFO 3244. The study showed that the same apparent molecular weights and same catalytic properties, but they were distinct from each other in subunit molecular mass.

The article discusses a study which investigated the energy metabolism and respiratory chain of Asaia bogorensis. A. bogorensis is a unique acetic acid bacterium with unusual properties including scanty acetic acid production and rapid growth. The found that A. bogorensis lacks quinoprotein alcohol dehydrogenase, but has sugar and sugar alcohol-oxidizing enzymes specific to the respiratory chain of Gluconobacter.

Pseudomonas putida HK5 produces three different quinoprotein alcohol dehydrogenases: ADH-I, ADH-IIB and ADH-IIG. Gene organization of qedA, the gene for ADH-I, and other 10 genes in the cluster was related to the genome sequences of five other Pseudomonas strains. Insertion mutations in either qedA, exaE or agmR eliminated ADH-I activity, although the mutants were still able to grow on ethanol but more slowly than the wild-type strain. Mutant analysis demonstrated the requirement of agmR and exaE in ADH-I expression, and the tentative involvement of agmR, but not exaE, in the induction of ADH-IIB and ADH-IIG activities. [ABSTRACT FROM AUTHOR]

The article reports on results of a study of physiological roles of two membrane-bound dehydrogenases responsible for d-sorbitol oxidation in Gluconobacter frateurii. A description of the study design and method is presented. The study concluded that pyroloquinoline quinone-dependent glycerol dehydrogenase (PQQ-GLDH) connects efficiently to cytochrome bo3 terminal oxidase.

The article reports on the results of a study of the occurrence of a novel NADH dehydrogenase (NADH-dh), distinct from the old yellow enzyme, in gluconobacter strains A description of the experimental set-up and measurement methods is presented. The study concluded that the N-terminal amino acid sequence of NADPH-dh matched a putative oxidoreductase containing the old yellow enzyme-like FMN-binding domain.

The article describes the preparation of two enzymes, 5-keto D-gluconate (5KGA) reductase (5KGR) and 2-keto D-gluconate (2KGA) reductase (2KGR) for quantification of 5KGA and 2KGA. According to the authors, the gene for 5KGR was found in the genome of Gluconobacter oxydans 621H and was identified as GOX2187, and the gene for 2KGR was identified as GOX0417 from the N-terminal amino acid sequence of the partially purified enzyme. The overexpression of 5KGR and 2KGR in Escherichia coli was noted.

The article examines the purification and properties of NADP-dependent shikimate dehydrogenase (SKDH) from Gluconobacter oxydans IFO 3244 and its application to enzymatic shikimate production. It showed a single protein band on native-polyacrylamide gel electrophoresis accompanying activity when SKDH was purified from Gluconobacter oxydans IFO 3244. It required NADP exclusively and catalyzed only the shuttle reaction between shikimate and 3-dehydroshikimate.

The article presents a study on the high shikimate (SKA) production from quinate with two enzymatic systems of acetic acid bacteria. 3-Dehydroshikimate was formed with a yield of 57 to 77 percent from quinate thru 3-dehydroquinate by two successive enzyme reactions, quinoprotein quinate dehydrogenase and 3-dehydroquinate dehydratase, in the cytoplasmic membranes of acetic acid bacteria. A strategy for high SKA production was proposed based on the data presented.

The article discusses the study about the conversion of capsular polysaccharide (CPS) to extracellular polysaccharide (EPS) by galE deletion in acetobacter tropicalis. A clone of the galE gene from Acetobacter tropicalis was performed. Results show that the cloned gene does not produce CPS, but secreted an EPS, which is purified and having a unique characteristic from the original CPS.

The article discusses the results of a study which examined membrane-bound NAD(P)-independent formaldehyde-oxidizing enzyme from Acetobacter sp. SKU 14. The enzyme was solubilized from the membrane fraction of glycerol-grown cells and was purified to homogeneity through column chromatographies. The purified enzyme was characterized as a quinoprotein alcohol dehydrogenase.

Examines the molecular properties of membrane-bound FAD-containing D-sorbitol dehydrogenase (SLDH) from thermotolerant Gluconobacter frateurii, an acetic acid bacteria. Industrial application of Gluconobacter strains; Role of SLDH in L-sorbose fermentation; Method used for the purification of FAD-SLDH from Gluconobacter frateurii.

Cites a study presenting the purification of type II NADH dehydrogenase of Corynebacterium glutamicum (NDH-2) from an ndh over-producing strain. Enrichment of NDH-2 by 6-fold higher specific activity of NADH after purification; Ability of the purified enzyme to catalyze electron transfer from NADH and NADPH to oxygen as well as various artificial quinone analogs; Presentation of C. glutamicum NDH-2 as a source of the reactive oxygen species, superoxide and hydrogen peroxide, concominant with NADH and NADPH oxidation.

Abstract: Quinoprotein quinate dehydrogenase (QDH) is a membrane-bound enzyme containing pyrroloquinoline quinone (PQQ) as the prosthetic group. QDH in Gluconobacter oxydans IFO3244 was found to be inducible by quinate and it is not constitutively expressed in the absence of quinate. The purification of holo-form of QDH to nearly homogeneity was achieved. The purified QDH appears to have two subunits of approximately 65 and 21 kDa, which could be the result of proteolysis of single polypeptide. Kinetic analysis indicated that the purified enzyme is much more specific to quinate than QDH from Acinetobacter calcoaceticus. The efficiency of the artificial electron acceptor was also determined. [Copyright &y& Elsevier]

The function of type II NADH dehydrogenase (NDH-2) in Gram-positiveCorynebacterium glutamicumwas investigated by preparing strains withndh, the NDH-2 gene, disrupted and over-expressed. Although disruption showed no growth defects on glucose minimum medium, the growth rate of the over-expressed strain was lower compared with its parent,C. glutamicumKY9714.Ndh-disruption and over-expression did not lead to a large change in the respiratory chain and energetics, including the cytochrome components and the H+/O ratio. However, in the strain that lacked NDH-2, membranel-lactate oxidase activity increased, while NDH-2 over-expression led to decreasedl-lactate and malate oxidase activities. In addition, relatively high cytoplasmic lactate dehydrogenase (LDH) activity was always present as was malate dehydrogenase, irrespective of NDH-2 level. Furthermore,l-lactate or malate-dependent NADH oxidase activity could be reproduced by reconstitution with the membranes and the cytoplasmic fraction isolated from the disruptant. These results suggest that coupling of LDH and the membranel-lactate oxidase system, together with the malate-dependent NADH oxidase system, operates to oxidize NADH when the NDH-2 function is defective inC. glutamicum. [ABSTRACT FROM AUTHOR]

Discusses 3-dehydroquinate production from quinate by oxidative fermentation with Gluconobacter strains of acetic acid bacteria and the further conversion of 3-dehydroquinate to the intermediates in the shikimate pathway. Quinate oxidation by acetic acid bacteria; Quinate oxidation by immobilized cells of acetic acid bacteria; Chromatographic detection of oxidative products.

Discusses the purification and characterization of membrane-bound quinoprotein quinate dehydrogenase (QDH). Occurrence of QDH in aerobic bacteria; Titration of apo-QDH with PQQ; Properties of purified QDH; Identification of quinate oxidation product by the membrane-bound QDH.

Amine oxidase AO-I from Aspergillus niger AKU 3302 has been reported to contain topa quinone (TPQ) as a cofactor; however, analysis of the p-nitro-phenylhydrazine-derivatized enzyme and purified active site peptides showed the presence of a carboxylate ester linkage of TPQ to a glutamate. The catalytic functionality of such a cross-linked cofactor has recently been shown unlikely by spectroscopic and voltammetric studies on synthesized model compounds. We have obtained resonance Raman spectra of native and substrate-reduced AO-I demonstrating that the catalytically active cofactor is unmodified TPQ. The primary structure of the enzyme (GenBank acc. no. U31869) has been reviewed and updated by repeated isolation and sequencing of AO-I cDNA. This allowed rectification of several errors that account for previously reported low homology to other amine oxidases in the regions around copper binding histididyl residues. The results were confirmed by cloning the ao-1 structural gene (GenBank acc. no. AF362473). Analysis of the gene 5'-upstream region of the gene revealed potential binding sites for an analog of NIT2, the nitrogen metabolism regulatory protein found in Neurospora crassa and other fungi. The molecular structure of AO-I was modeled by a comparative method using published crystal structures of amine oxidases as templates. [ABSTRACT FROM AUTHOR]

Investigates the molecular cloning of quinohemoprotein alcohol dehydrogenase (ADH IIB) from Pseudomonas putida HK5. Identification of a gene diverntly transcribed against qbdBA with NAD-dependent adelhyde dehydrogenase.

Describes the purification and characterization of two NAD-dependent alcohol dehydrogenase (ADH) from the soluble fraction of Acetobacter pasteurianus CN5-2. Homology of the ADH I and ADH II to the long-chain and short-chain ADH families; Difference in their kinetics on ethanol, acetaldehyde, NAD and NADH.

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. [ABSTRACT FROM AUTHOR]