Acetobacter methanolicus sp. nov., an Acidophilic Facultatively Methylotrophic Bacterium

Uhlig, Hilde; Karbaum, Klaus; Steudel, A.

doi:10.1099/00207713-36-2-317

Cited by 77 publications

(50 citation statements)

References 4 publications

(4 reference statements)

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“…With the exception of Acetobacter methanolicus (Uhlig et al, 1986), all known Gram-negative methanolutilizing bacteria are obligate methylotrophs, growing only on methanol and sometimes methylamine. Originally described under various names, they are now (Urakami & Komagata, 1986) and Methylophilus (Jenkins et al, 1987).…”

Section: Introductionmentioning

confidence: 99%

Isolation and phenotypic characterization of methanol oxidation mutants of the restricted facultative methylotroph Methylophaga marina

Janvier

Frank

Luttik

et al. 1992

Journal of General Microbiology

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Methanol oxidation in Mefhyfophuga marina appears to be organized in a way similar to that in other Gramnegative methylotrophs, involving a quinoprotein methanol dehydrogenase (MDH; EC 1.1.99.8) and two soluble cytochromes. The MDH is composed of two different subunits, most probably arranged in an a2P2 structure. The two cytochromes are of the e-type and differ in size (molecular mass 19.5 and 12.5 kDa) and isoelectric point (PI 4.6 and 9.2). The one with the lowest isoelectric point, commonly designated as cytochrome eL, is able to oxidize reduced MDH. Taking advantage of the ability of M. marina, a restricted facultative methylotroph, to utilize fructose as a growth substrate, mutants impaired in methanol utilization were isolated after application of optimal concentrations of ethylmethane sulphonate. Three classes of methanol oxidation mutants were obtained. Class I mutants were affected in a global regulation of the synthesis of both apo-methanol-dehydrogenase and cytochrome cL as well as PQQ (pyrroloquinoline quinone). Class I1 mutants did not produce an active MDH, but instead a comparable amount of a 65 kDa protein was found in the cell-free extract upon SDS-PAGE. This mutant protein was purified and compared to wild-type MDH. It was located in the periplasm, but unlike MDH it was composed of only two identical large subunits. Each of these subunits was able to bind one molecule of PQQ. An antiserum raised against wild-type MDH did not react with the mutant protein. Conversely, an antiserum raised against mutant protein weakly cross-reacted with wild-type MDH, suggesting that the presence of the P-subunit in MDH dramatically changes its immunochemical behaviour. Three of the class I1 mutants did not produce PQQ. In the presence of PQQ, partial revertants able to grow on methanol medium were obtained (class 111). Class 111 mutants produced a stable apo-MDH consisting of a-and P-subunits and showing a normal reaction with the anti-MDH serum. Although this apo-MDH can bind PQQ, enzymic activity was not restored in nitro. This suggests that, in addition to apo-MDH and PQQ, other factors are required for the assembly of an enzymically active MDH.

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Section: Introductionmentioning

confidence: 99%

Isolation and phenotypic characterization of methanol oxidation mutants of the restricted facultative methylotroph Methylophaga marina

Janvier

Frank

Luttik

et al. 1992

Journal of General Microbiology

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“…It was the kind gift of Professor W. Babel (Institut fur Biotechnologie, Leipzig, Germany). Stock cultures were maintained on methanol-containing minimal agar slopes at 4 "C or in 30% (v/v) glycerol at -20 "C. The defined growth medium was that described by Uhlig et al (1986), containing methanol (1 %, v/v).…”

Section: Methodsmentioning

confidence: 99%

The o-type oxidase of the acidophilic methylotroph Acetobacter methanolicus

Chan

Anthony

1991

Journal of General Microbiology

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Membranes of the acidophilic methylotroph Acetobucter methanolicus contained only b-and c-type cytochrome and a CO-binding b-type cytochrome. An azide-sensitive oxidase that oxidizes cytochrome c and ascorbatel TMPD was solubilized from the membrane with a mixture of CHAPS and Zwittergent3-12 (1.7 fold increase in specific activity with 32% yield). The solubilized oxidase is unusually stable with respect to high ionic strength (200 mM-NaC1) and stable between pH 4.0 and 6.8. Of the two soluble c-type cytochromes from A. methanoficus only the typical class I cytochrome (cytochrome cH) was a good substrate, as was equine cytochrome c. The oxidase was partially purified by anion-exchange chromatography but further purification proved impossible. The yield with respect to equine cytochrome c oxidation was l8%, with a 22-fold purification, but during purification most of the activity with respect to cytochrome cH and TMPD was lost. Neutral phospholipids had little effect on activity of the oxidase but the charged phospholipids phosphatidylglycerol and phosphatidylserine stimulated activity up to about fourfold. It proved impossible to incorporate the oxidase into active lipoprotein vesicles. During the purification process the pH optimum for oxidation of cytochrome cH was unchanged (pH 5.6) whereas that for oxidation of equine cytochrome changed from pH 9.5 to 7.5 and the sensitivity of the oxidase to azide changed from non-competitive to competitive during the purification process. The partially-purified oxidase contained only btype cytochrome, some of which was CO-reactive. It is proposed that the oxidase is a cytochrome co type of oxidase that loses its cytochrome c component during the purification process and is only able to oxidize c-type cytochromes if these can be formed into a 'reconstituted' cytochrome co with the partially-purified oxidase. IntroductionThe present paper describes work on an unusual o-type oxidase from the acidophilic methylotroph Acetobacter methanolicus, which grows at pH 4 on methanol as its sole carbon and energy source (Steudel & Babel, 1982;Steudel et al., 1980). As in other methylotrophic bacteria, methanol is oxidized by way of the quinoprotein methanol dehydrogenase, its specific electron acceptor cytochrome cL, and a typical class I c-type cytochrome which is oxidized by a membrane oxidase (Anthony, 1982(Anthony, , 1986Elliott & Anthony, 1988). Except for the oxidase all these proteins are periplasmic and so operate at pH4, the pH of the growth medium. Preliminary observations suggested that this oxidase may be more stable to extremes of pH and salt concentration compared with cytochrome oxidases in other bacteria.The bacterial o-type oxidases are defined loosely as oxidases having a CO-binding cytochrome b, which for convenience is usually referred to as cytochrome o and which is assumed to be the oxygen-reactive site. Cytochrome o was first described in Staphylococcus aureus (Chance, 1953 ;Castor & Chance, 1959) and the 0-type oxidases have since been shown to be the most widely...

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“…All experiments were carried out with the bacterial strain .4. methanolicus MB 58/4 and its phage Acre1, both isolated and characterized in the Institute of Biotechnology, Leipzig (Wfinsche et aL, 1983a;Uhlig et al, 1986). Amino acid-auxotrophic mutants were obtained by a combination of u.v.-irradiation and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) treatment of bacterial suspensions under standardized conditions.…”

Section: Methodsmentioning

confidence: 99%

Phage Acm1-mediated transduction in the facultatively methanol-utilizing Acetobacter methanolicus MB 58/4

Kiesel

Wünsche

1993

Journal of General Virology

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Acetobacter methanolicus sp. nov., an Acidophilic Facultatively Methylotrophic Bacterium

Cited by 77 publications

References 4 publications

Isolation and phenotypic characterization of methanol oxidation mutants of the restricted facultative methylotroph Methylophaga marina

Isolation and phenotypic characterization of methanol oxidation mutants of the restricted facultative methylotroph Methylophaga marina

The o-type oxidase of the acidophilic methylotroph Acetobacter methanolicus

Phage Acm1-mediated transduction in the facultatively methanol-utilizing Acetobacter methanolicus MB 58/4

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