Cobamide Coenzymes Research Articles

The effect of chemical modifications of cobamide co-enzyme on the rate of substitution of trans-axial ligands

The effect of aglycone substitution in the 5′-desoxynucleoside part of cobamide co-enzyme 1 1 Notation : cobamide co-enzyme [α(5,6-dimethylbenzimidazolyl)-Co-5′-desoxyadenosylcobamide, DBC]. , modifications in the sugar part of desoxynucleoside ligand, the effect of desoxyglycosyl ligands and modifications in the “lowel” axial position on the stability of CoC and CoN Bz bonds in the cyanide cleavage reactions have been studied. Cyanide reactions involving octahedral derivatives of cobamide co-enzymes proceed via two steps: a fast step ( k 1) corresponding to the substitution of the “lower” 5,6-dimethylbenzimidazole ligand, and a slow step ( k 2) corresponding to the simultaneous cleavage of the CoC and Nglycoside bonds. Other mechanisms are involved in cyanide reactions with analogues modified at the “lower” axial ligand which presumably possess a similar structure to penta-coordinated complexes.

Biosynthesis and properties of 2-thioadenyl cobamide and its coenzyme form

Barker et al. [l-2] first isolated adenyl cobamide coenzyme and subsequently cobalamin coenzyme from cultures of Clostridium tetanomorphum and later from Propionibacterium sp. Thereafter, many reports have appeared on the roles of the coenzyme forms of cobalamin in a number of important biochemical reactions. Kamikubo et al. [3] reported that biosynthetic 2-chloroadenyl cobamide coenzyme from growing Propionibacterium arabinosum cells was active as a coenzyme for propanediol dehydrase. However, no further reports have appeared on the biochemical functions of cobamide coenzymes with purines in the nucleotide moiety. In this paper, the biosyhthesis of 2-thioadenyl cobamide and its coenzyme form using resting Pr. arabinosum cells, their physico-chemical properties and biological as well as biochemical activities are dealt with.

Cobamides and Ribonucleotide Reduction: V. A KINETIC STUDY OF THE RIBONUCLEOSIDE TRIPHOSPHATE REDUCTASE OF LACTOBACILLUS LEICHMANNII

Abstract Initial velocities of the reaction catalyzed by the ribonucleotide reductase of Lactobacillus leichmannii have been measured spectrophotometrically in the presence of various combinations of cobamide coenzymes, ribonucleoside triphosphate substrates, deoxyribonucleoside triphosphate activators and the reduced nicotinamide adenine dinucleotide phosphate-linked thioredoxin system of Escherichia coli which acted as the reductant. Of the ribonucleotide substrates tested, guanosine triphosphate is reduced most rapidly and its reduction is not stimulated significantly by the addition of deoxyribonucleotides. The reduction rate of adenosine triphosphate is elevated to that of GTP by the addition of dGTP; similarly, the rate of cytosine triphosphate reduction is increased to this rate by dATP. Smaller increases were observed in the rates of reduction of uridine triphosphate (with dCTP) and of inosine triphosphate (with deoxythymidine triphosphate). This activation by deoxyribonucleotides may be related to a decrease which they cause in the apparent Michaelis constant for deoxyadenosylcobalamin coenzyme. In the absence of added deoxyribonucleotides the apparent Michaelis constant for deoxyadenosylcobalamin is dependent upon the nature and concentration of the ribonucleotide substrate used in the reaction mixture. Nonlinear plots of reciprocal velocity against reciprocal ribonucleotide concentration and against reciprocal deoxyadenosylcobalamin concentration are obtained under certain conditions. The interdependence of coenzyme, substrate, and activator is interpreted as indicating that the enzyme has an allosteric site and that occupation of this site by a deoxyribonucleoside- or ribonucleoside-triphosphate increases the activity of the enzyme. The activation is believed to be effected at least in part via an increase in affinity of the enzyme for deoxyadenosylcobalamin. 5,6-Dimethylbenzimidazolyl-5'-deoxyadenosylcobamide (5'-deoxyadenosylcobalamin) and benzimidazolyl-5'-deoxyadenosylcobamide were equally active as coenzymes while adenosyl-5'-deoxyadenosylcobamide and the anilide of 5'-deoxyadenosylcobalamin, though active, gave apparent Km values 3 to 4 times greater than the former two coenzymes. 2',5'-Dideoxyadenosylcobalamin showed very slight activity and acted as a competitive inhibitor of 5'-deoxyadenosylcobalamin, whereas 5'-deoxycytidylcobalamin was inactive and a poor inhibitor.

Book Review: “Comprehensive Biochemistry, Vol. 16: Hydrolytic Reactions; Cobamide and Biotin Coenzymes”

Book Review: “Comprehensive Biochemistry, Vol. 16: Hydrolytic Reactions; Cobamide and Biotin Coenzymes”

Effect of Cobalt on the Non-nodulated Legume

IT has been demonstrated on several occasions that the addition of small quantities of cobalt as a nutrient has a stimulating effect on the growth of nodulated legumes1–3, whether they are grown in water culture, in sand culture or in the field. This has generally been attributed to an effect on the symbiotic fixation of nitrogen by the nodule, and the discovery that cobamide co-enzymes are present in the nodules of several leguminous species, as well as in the nodules of certain non-legumes4, enhances the probability of this hypothesis.

The Relationship of Cobalt Requirement to Propionate Metabolism in Rhizobium

Cobalt is known to be essential for symbiotic nitrogen fixation by legumes (l-5), but a requirement of the element for growth of leguminous plants supplied with adequate fixed nitrogen has not been shown. Ahmed and Evans (3) provided evidence that cobalt was required for the bacteria within the nodules of inoculated legumes but was not required for the leguminous plants per se. Lowe, Evans, and Ahmed (6) and Lowe and Evans (7) subsequently showed that cobalt is necessary for normal growth of five species of Rhixobium. More recently, Kliewer and Evans (8-10) analyzed nodules of a variety of symbionts and cells from several species of Rhizobium and observed the presence of cobamide coenzymes in each of the organisms. In addition, they (9) isolated and identified the dimethylbenaimidazolylcobamide coenzyme in extracts of cells isolated from soybean nodules and in cells of a pure culture of Rhizobium meliloti and presented data (10) showing that cobalt deficiency not only limited the growth of Rhizobium japonicum and R. meliloti but also limited the synthesis of cobamide coenzymes in these organisms. The Bn coenzyme has been shown to be involved in propionate metabolism in animals (11-13) and in Propionibacterium sherma&i (14, 15). In addition, Arnstein and White (16) have shown that Bn deficiency in Ochromonas malhamensis resulted in a reduced capacity of the organism to oxidize propionate. Ayers (17) has described similar effects of Blz deficiency in a Flavobacterium species. On the basis of these reports, experiments were conducted to determine whether Rhizobium would metabolize propionate and the possible effect of cobalt deficiency on propionate utilization in these bacteria. In a preliminary communication (18), evidence was presented showing that the rate of oxidation of propionate by cell suspensions of R. meliloti was strikingly affected by the concentration of cobalt in the culture medium. In contrast, the oxidation of glutamate was not affected by cobalt deficiency. In this paper, evidence is presented showing that enzymes necessary for the catalysis of the conversion of propionate to succinyl coenzyme A by a pathway involving methylmalonyl coenzyme A are widely distributed in Rhizobium and that cobalt deficiency in R. meliloti results in insufficient Blz coenzyme for the function of methylmalonyl coenzyme A mutase in cell-free extracts.

STUDIES OF THE MECHANISM OF ACTION OF COBAMIDE COENZYMES.

Annals of the New York Academy of SciencesVolume 112, Issue 2 p. 695-702 STUDIES OF THE MECHANISM OF ACTION OF COBAMIDE COENZYMES R. H. Abeles, R. H. Abeles University of Michigan Medical School, Ann Arbor, Mich.Search for more papers by this authorH. A. Lee Jr., H. A. Lee Jr. University of Michigan Medical School, Ann Arbor, Mich.Search for more papers by this author R. H. Abeles, R. H. Abeles University of Michigan Medical School, Ann Arbor, Mich.Search for more papers by this authorH. A. Lee Jr., H. A. Lee Jr. University of Michigan Medical School, Ann Arbor, Mich.Search for more papers by this author First published: April 1964 https://doi.org/10.1111/j.1749-6632.1964.tb45046.xCitations: 25AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume112, Issue2Vitamin B12 CoenzymesApril 1964Pages 695-702 RelatedInformation

FACTORS AFFECTING THE FORMATION OF COBAMIDE COENZYMES IN CLOSTRIDIUM TETANOMORPHUM.

Toohey, J. I. (University of California, Berkeley), and H. A. Barker. Factors affecting the formation of cobamide coenzymes in Clostridium tetanomorphum. J. Bacteriol. 87:504-509. 1964.-Tests were carried out to determine the optimal culture conditions for the production of cobamide coenzymes in Clostridium tetanomorphum strain H1. A method is described for carrying out coenzyme determinations on the cells from 10-ml cultures of the bacterium. In a basal medium containing magnesium sulfate, ferrous sulfate, manganese sulfate, sodium molybdate, calcium chloride, and potassium phosphate, the optimal concentration of monosodium glutamate was 0.1 m and of yeast extract was 3 g per liter. Addition of glucose at a concentration of 0.05 m was found to double the yield of cells and to increase tenfold the specific coenzyme yield. Addition of cobaltous chloride (2 x 10(-5)m) also increased coenzyme production. Addition of benzimidazole caused an apparent increase in coenzyme production by causing the synthesis of the highly active benzimidazole analogue. Addition of methionine (5 x 10(-6)m) appeared to inhibit coenzyme production.

Identification of Cobamide Coenzyme in Nodules of Symbionts & Isolation of the B12 Coenzyme From Rhizobium meliloti

Recently, Barker et al. (5) have isolated from Clostridium tetanomorphum and Propionibacterium shermanii three highly purified cobamide coenzymes. These were subsequently identified as adenylcobamide (AC), benzimidazolylcobamide (BC), and 5,6-dimethylbenzimidazolylcobamide (DBC). It has been shown that these compounds function as cofactors in the conversion of glutamate to ,-methyl aspartate (4), in the isomeration of methylmalonyl CoA to succinyl CoA (9), and in the conversion of 1,2-propanediol to propionaldehyde (1). All of the cobamide coenzyme molecules contain cobalt, but they differ in that adenine, benzimidazole, or dimethylbenzimadazole bases are constituents of the AC, BC, and DBC coenzymes, respectively (4, 5). It has been established that several Rhizobium species (7,8) and legumes (2,3) grown without a fixed source of nitrogen, require cobalt for normal growth. Evidence also has been provided that Rhizobium meliloti and nodules from several legumes and alder, contain a cobamide coenzyme (6)3 capable of serving as a cofactor for the enzymatic conversion of glutamate to P-methyl aspartate. Identification and characterization of the cobamide coenzymes in these organisms is obviously of interest. A preliminary paper reports that the B12 coenzyme isolated from Rhizobium meliloti was the 5,6-dimethylbenzimidazolylcobamide compound (6). This paper presents evidence establishing the identity of the B12 coenzyme from R. meliloti. Also, data on the distribution of B12 coenzymes in the nodules of symbionts are reported.

THE INFLUENCE OF COBAMIDES ON THE ENDOGENOUS AND EXOGENOUS RESPIRATION OF A MARINE BACTERIUM

The oxidation of propionate, isobutyrate, valerate, isoleucine, and valine as well as the endogenous respiration of a B12-dencient marine bacterium was stimulated by the addition of vitamin B12to washed cell suspensions. The B12analog, 2-methyladenylcobamide cyanide, also stimulated endogenous respiration, but cobinamide was inactive. During autorespiration ammonia was continuously evolved by the cells indicating the presence of a nitrogenous endogenous substrate. When vitamin B12was supplied to the cells, the ammonia evolved was decreased, while the rate of oxygen uptake was increased. The possible involvement of cobamide coenzymes is discussed in relation to these findings.

The Enzymatic Synthesis of Cobamide Coenzymes

The Enzymatic Synthesis of Cobamide Coenzymes

A cobamide-requiring glycerol dehydrase from an acrolein-forming lactobacillus

A glycerol dehydrase has been isolated from Lactobacillus 208-A which requires a cobamide coenzyme and a univalent cation for activity. The enzyme is measured by the colorimetric determination of the acrolein formed from β-hydroxypropionaldehyde produced from glycerol by the dehydrase. Vitamin B 12 inhibits the dehydrase. The degree of inhibition by vitamin B 12 is greater when it contacts the apoenzyme before the cobamide coenzyme. The toxicity of mercurial compounds can be relieved by cobamide coenzymes or by 2,3-dimercaptopropanol (BAL). This dehydrase has an optimum pH of 5.8–6.0.

Derivatives of 1,4-dioxo-cyclohexane as possible intermediates in isomeric changes dependent on cobamide coenzymes.

Derivatives of 1,4-Dioxo-Cyclohexane as Possible Intermediates in Isomeric Changes Dependent on Cobamide Coenzymes

Structure of Cobamide Coenzymes: Influence of pH on Absorption Spectra and Ionophoretic Mobilities

Structure of Cobamide Coenzymes: Influence of pH on Absorption Spectra and Ionophoretic Mobilities

Purification and Properties of Cobamide Coenzymes Obtained from Propionibacterium arabinosum

Purification and Properties of Cobamide Coenzymes Obtained from Propionibacterium arabinosum

The enzymatic synthesis of cobamide coenzymes

The enzymatic synthesis of cobamide coenzymes

Detection of cobamide coenzymes in microorganisms by the ionophoretic-bioautographic method

The ionophoretic-bioautographic method of detecting cobinamide and cobamide vitamins in millimicrogram quantities has been modified and applied to the detection of cobamide coenzymes in microbial extracts. For the detection of cobamide coenzymes, the cells must be extracted with a neutral buffer in the dark. When cells are extracted with either an alkaline or acid cyanide solution or when the extract is exposed to light, only the cobamide vitamins and other coenzyme decomposition products are observed. Cobamides with the ionophoretic mobility of the cobamide coenzymes have been shown to be present in extracts of Escherichia coli B, E. coli 113-3, Bacillus megatherium, Streptomyces fradiae, Propionibacterium arabinosum, P. shermanii, and Clostridium tetanomorphum. The coenzymes present in several of these organisms have been specifically identified. E. coli 113-3 was shown to convert vitamin B 12 to the 5,6-dimethylbenzimidazolylcobamide coenzyme (coenzyme B 12). When grown with cobinamide and benzimidazole, the benzimidazolylcobamide coenzyme was formed.

Isolation of Coenzyme B12 from Liver

The identity of the major cobamide in liver has not been satisfactorily established. Cyanocobalamin (vitamin B12, 5,6dimethylbenzimidazolylcobamide cyanide), which has been obtained in crystalline form with liver as a source material (1, 2), is certainly a very minor component of the cobalamins in liver. Analyses of liver concentrates, both by a paper chromatographicbioautographic method (3) and by a chemical method (4), have shown that cyanocobalamin is usually not present in detectable amounts and seldom constitutes as much as 10% of the total cobalamin. The preparation of cyanocobalamin from liver probably was made possible by the presence of cyanide in the charcoal used in the isolation process. Examination of a liver extract by the chromatographic-bioautographic method (3) indicated that hydroxocobalamin (vitamin B12b, 5,6-dimethylbenzimidazolylcobamide hydroxide) was the most abundant cobalamin. This compound was also obtained in pure form from liver (1, 5). However, the recent discovery of the cobamide coenzymes in bacteria (6,7) and the demonstration that they constitute a large part of the total cobamides in these organisms (8,9) raised the possibility that coenzyme Bit (5,6-dimethylbenzimidazolylcobamide coenzyme, DMBC coenzyme, DBC coenzyme) may be a constituent of liver. Since coenzyme B12, like the adenylcobamide coenzyme (lo), is relatively unstable and is readily converted to hydroxocobalamin or cyanocobalamin by treatment with light or cyanide,’ respectively, the inadvertent decomposition of the coenzyme, if present in liver extracts, could account for previous observations on the occurrence of the two cobalamin vitamins. The present paper reports some observations on the occurrence and abundance of a cobamide coenzyme, which has been identified as coenzyme Blz, in livers of rabbit, chicken, sheep, and man. A brief report of the occurrence of coenzyme Blz in rabbit liver has already appeared (7).

Observations on the structure of cobamide coenzymes

Observations on the structure of cobamide coenzymes

Isolation and Properties of Crystalline Cobamide Coenzymes Containing Benzimidazole or 5,6-Dimethylbenzimidazole

Isolation and Properties of Crystalline Cobamide Coenzymes Containing Benzimidazole or 5,6-Dimethylbenzimidazole