Purification and properties of vitamin B12-dependent ribonucleotide reductase from Rhizobium meliloti.

Inukai, Shinji; Sato, Kazuyoshi; Shimizu, Sakayu

doi:10.1271/bbb1961.43.637

Cited by 6 publications

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“…A similar approach, based on the vitamin B12 requirement in anaerobiosis for the growth of an S. typhimurium metE mutant, has been described to screen Cob mutants (29). This type of screening can probably be used for bacteria such as R. meliloti, the growth of which depends on the presence of vitamin B12 in the medium when cobalt ions are absent (27,28). In this case, the vitamin B12-dependent enzymes involved are a ribonucleotide reductase (28) and a methionine synthetase (27).…”

Section: Discussionmentioning

confidence: 99%

Cloning and analysis of genes involved in coenzyme B12 biosynthesis in Pseudomonas denitrificans

Cameron¹,

Briggs²,

Pridmore³

et al. 1989

J Bacteriol

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Cobalamin synthesis probably requires 20 to 30 different enzymatic steps. Pseudomonas putida and Agrobacterium tumefaciens mutants deficient in cobalamin synthesis (Cob mutants) have been isolated. In P. putida, Cob mutants were identified as being unable to use ethanolamine as a source of nitrogen in the absence of added cobalamin (deamination of ethanolamine requires coenzyme B12 as a cofactor). In A. tumefaciens, Cob mutants were simply screened for their reduced cobalamin synthesis. A genomic library of Pseudomonas denitrificans was constructed on a mobilizable wide-host-range vector. Eleven plasmids from this library were able to complement most of these mutants. By complementation and restriction mapping analysis, four genomic loci of P. denitrificans were found to be responsible for complementation of the Cob mutants. By subcloning fragments from the four genomic loci, we identified at least 14 different genes involved in cobalamin synthesis.Cobalamins are among the most complex nonpolymeric natural products known (4). The coenzyme B12 biosynthetic pathway contains the following steps ( Fig. 1): (i) formation of uroporphyrinogen III, the common intermediate for the synthesis of hemes, chlorophylls, cobalamin, sirohemes, and F430 (23); (ii) conversion of uroporphyrinogen III into cobyrinic acid; (iii) formation of cobinamide from cobyrinic acid; and (iv) conversion of cobinamide into coenzyme B12 (for reviews on cobalamin biosynthesis, see references 3, 22, 26, 41, and 46). Uroporphyrinogen III is synthesized in four distinct and well-known enzymatic steps from succinyl coenzyme A and glycine (3). In contrast, the biosynthetic pathway from uroporphyrinogen III to coenzyme B12 is not clearly understood, since only one enzymatic activity has been partially purified (38) and the precise number of enzymatic steps is not known. In addition, the intermediates between porphobilinogen and cobyrinic acid are very unstable and sensitive to oxygen (3).

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Section: Discussionmentioning

confidence: 99%

Cloning and analysis of genes involved in coenzyme B12 biosynthesis in Pseudomonas denitrificans

Cameron¹,

Briggs²,

Pridmore³

et al. 1989

J Bacteriol

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“…Vitamin B 12 is also required for the activity of a B 12 -dependent ribonucleotide reductase found in rhizobia, Lactobacillus leichmannii, and several other bacteria, but not in E. coli (12). In the presence of methionine alone (i.e., without cobalt), the ribonucleotide reductase was found to be synthesized at higher levels in S. meliloti cells.…”

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confidence: 96%

“…The most efficient version of this enzyme requires vitamin B 12 as a cofactor (13,17). In bacteria the requirement for cobalt is primarily due to its presence in vitamin B 12 and the important role of this vitamin in methionine biosynthesis. It was found that cobalt (cobalt chloride, 5 ng/ml) could be substituted for methionine to permit growth of S. meliloti strains in minimal medium (Fig.…”

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confidence: 99%

“…In the presence of methionine alone (i.e., without cobalt), the ribonucleotide reductase was found to be synthesized at higher levels in S. meliloti cells. This was reported to affect cell morphology but not growth (12). For this reason it is probably preferable to always supplement the cells with cobalt.…”

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“…Methionine can also be synthesized in many bacteria by a second homocysteine methyltransferase induced in the absence of cobalt, which does not require vitamin B 12 , but this enzyme is extremely inefficient (13,17). We tested for slow growth of S. meliloti cells in minimal medium containing biotin but neither cobalt nor methionine during extended incubations (Fig.…”

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Sinorhizobium meliloti Cells Require Biotin and either Cobalt or Methionine for Growth

Watson

Heys

Martin

et al. 2001

Appl Environ Microbiol

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Sinorhizobium meliloti is usually cultured in rich media containing yeast extract. It has been suggested that some components of yeast extract are also required for growth in minimal medium. We tested 27 strains of this bacterium and found that none were able to grow in minimal medium when methods to limit carryover of yeast extract were used during inoculation. By fractionation of yeast extract, two required growth factors were identified. Biotin was found to be absolutely required for growth, whereas previously the need for this vitamin was considered to be strain specific. All strains also required supplementation with cobalt or methionine, consistent with the requirement for a vitamin B 12 -dependent homocysteine methyltransferase for methionine biosynthesis.

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The Nature of Distribution of Ribonucleotide Reductases

Follmann

Harder

Hogenkamp

1988

Metabolism and Enzymology of Nucleic Acids

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Purification and properties of vitamin B12-dependent ribonucleotide reductase from Rhizobium meliloti.

Abstract: Ribonucleotide reductase from Rhizobium meliloti grown on the cobalt-deficient medium containing methionine was purified about 37-fold by streptomycin treatment, ammonium

Cited by 6 publications

References 0 publications

Cloning and analysis of genes involved in coenzyme B12 biosynthesis in Pseudomonas denitrificans

Cloning and analysis of genes involved in coenzyme B12 biosynthesis in Pseudomonas denitrificans

Sinorhizobium meliloti Cells Require Biotin and either Cobalt or Methionine for Growth

The Nature of Distribution of Ribonucleotide Reductases

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