Cobamide Coenzyme Research Articles

Effects of cobalt on plants

Effects of cobalt on plants

Aerobic degradation of choline by Proteus mirabilis: enzymatic requirements and pathway.

Cleavage of choline to trimethylamine and acetaldehyde by extracts of Proteus mirabilis requires both particulate and soluble protein fractions, K+, and a bound divalent metal cation. The reaction shows a long lag period, abolished only by preincubation of the particulate fraction in the complete reaction system. The two-carbon fragment produced is acetaldehyde; choline cleavage appears to be tightly coupled to dismutation of the acetaldehyde to ethanol and acetate, as indicated by stimulation by NAD+, ADP, and Fe2+ and inhibition by reagents reacting with acetaldehyde. The system is thus similar to that previously described in anaerobes (Desulfovibrio, Clostridium). Attempts to demonstrate a cobamide coenzyme requirement (as in the similar ethanolamine ammonia-lyase reaction) were unsuccessful; the reaction was carried out by fractions devoid of vitamin B12 activity (not supporting growth of Lactobacillus leichmannii) and insensitive to light.

Diol metabolism and diol dehydratase in Clostridium glycolicum

Levels of the five enzymes involved in the fermentation of 1,2-ethanediol and 1,2-propanediol in the strictly anaerobic bacterium, Clostridium glycolicum, were investigated. All enzymes with the exception of the first enzyme in the pathway, diol dehydratase, were found to be constitutive, stable to exposure to oxygen, and present in the cytosol. Diol dehydratase was found to be extremely oxygen sensitive and strongly associated with the cell membrane. Treatment with ionic and nonionic detergents, butanol, phospholipase A 2, or osmotic shock procedures failed to solubilize any diol dehydratase activity. Limited proteolysis using subtilisin released small amounts of activity. Diol dehydratase was found to be specific for 1,2-ethanediol and 1,2-propanediol and required the addition of a reducing agent for maximal activity. The enzyme was strongly inhibited by low concentrations of EDTA, ethylene glycol bis(β-aminoethyl ether)- N,N,N′,N′-tetraacetic acid, o-phenanthroline, hydroxylamine, hydroxyurea, and sulfhydryl reagents. Addition of adenosylcobalamin or high levels of intrinsic factor did not affect the reaction rate. Irradiation with light also did not inhibit the enzyme activity. These results suggest that the catalytic mechanism of diol dehydratase from C. glycolicum does not involve a cobamide coenzyme.

23] β-methylaspartate-glutamate mutase from Clostridium tetanomorphum

Publisher Summary This chapter provides an overview of β-methylaspartate-glutamate mutase from Clostridium tetanomorphum. The conversion of L-glutamate to threo-30-methyl-L-aspartate by glutamate mutase is the initial reaction in the anaerobic degradation of glutamate by Clostridium tetanomorphum and other Clostridium species. Two readily separable proteins called component E (MW 128,000) and component S (MW 17,000), a cobamide coenzyme and a sulfhydryl compound, are required for activity. Component S is reversibly inactivated by oxygen and, therefore, must be reduced by a sulfhydryl compound for full activity. Two assay methods are described in the chapter: a spectrophotometric assay done in the presence of air and an anaerobic assay. The spectrophotometric assay, which is generally used for estimating mutase activity, is based on the conversion of glutamate to mesaconate by the coupled action of mutase and β-methylaspartase. The anaerobic assay is based upon the conversion of β-methylaspartate to glutamate and is applicable only to purified mutase preparations in which β-methylaspartase is either absent or fully inhibited by Ca 2+ .

Microbial Metabolism of Amino Alcohols. Control of Formation and Stability of Partially Purified Ethanolamine Ammonia-lyase in Escherichia coli

Induction of ethanolamine ammonia-lyase formation in Escherichia coli required both the ethanolamine and vitamin B12, and was gratuitous during growth on glycerol. Ethanolamine analogues inhibited enzyme activity and inhibited growth with ethanolamine as the the nitrogen source, but did not act as inducers. Enzyme formation was more rapid when ethanolamine was added to cultures containing vitamin B12 rather than the reverse. Enzyme formation was subject to catabolic repression, glucose and acetate being particularly effective. Chloramphenicol, I-aminopropan 2-01 and 1,3-diaminopropan-2-01 prevented enzyme induction. Ethanolamine ammonia-lyase, resolved from its cobamide coenzyme, was purified 35-fold. The apoenzyme was stable for several days in the presence of ethanolamine, dithiothreitol, glycerol and K+ ions. Enzyme formation therefore requires both substrate and cobamide coenzyme to be present simultaneously as inducers.

Isolation and Identification of a Cobamide Coenzyme from the Tapeworm Spirometra mansonoides

A light-sensitive vitamin B12 derivative has been extracted from the adult cestode, Spirometra mansonoides. This corrinoid was identified as the cobamide coenzyme, adenosylcobalamin, by its chromatographic, chemical, and spectral properties.

Studies on methanol metabolism. III. Isolation of 5,6-dimethylbenzimidazolyl cobamide coenzyme from a methanol utilizing bacterium, Klebsiella sp. No. 101.

The isolation and identification of vitamin B12 produced in the cells of Klebsiella sp. No. 101 grown in a medium containing methanol as a sole carbon source were investigated. The B12-vitamers formed in the cells of the bacterium were extracted with hot ehtanol and purified by solvents extraction and ion-exchange cellulose column chromatography. The identity of the main component with 5,6-dimethylbenzimidazolyl cobamide coenzyme (DBCC) was demonstrated by paper chromatography, paper ionophoresis, absorption spectrophotometry of its coenzyme form, cyano form and nucleotide.

Demonstration of cobamide coenzyme in Ascaris suum

1. 1. Cobamide coenzyme was extracted from the body wall of adult female Ascaris suum maintained in Ellison's medium containing vitamin B 12. 2. 2. The identification of the coenzyme in purified fractions was based on (a) specific spectral changes following treatment of the preparation with light and cyanide, (b) coenzyme activity in the glutamate mutase reaction and (c) inactivation of the coenzyme activity by light. 3. 3. A fivefold increase in coenzyme occurred in worms maintained in Ellison's medium plus vitamin B 12, compared to the endogenous value. The data indicate that vitamin B 12 is converted by A. suum into the metabolically active coenzyme.

Affinity Specificity of Cobamide Coenzyme Analogues Containing Purine Derivatives for Glutamate Mutase

Affinity Specificity of Cobamide Coenzyme Analogues Containing Purine Derivatives for Glutamate Mutase

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.

Nicotinic Acid Metabolism: VI. PURIFICATION AND PROPERTIES OF α-METHYLENEGLUTARATE MUTASE (B12-DEPENDENT) AND METHYLITACONATE ISOMERASE

Abstract The conversion of α-methyleneglutarate (αMG) to dimethylmaleate (DMM) via methylitaconate (MeIT) is the reaction sequence that produces a symmetrical intermediate in the fermentation of nicotinic acid by Clostridium barkeri. αMG mutase catalyzes the isomerization of αMG in a B12 coenzyme-dependent reaction to form MeIT which in turn is isomerized to DMM by MeIT isomerase. Both enzymes are induced by growth on nicotinic acid and are not detectable in cells grown on glucose. The isomerase reaction favors the formation of DMM, whereas the mutase reaction favors the formation of αMG. Both reactions were demonstrated to be reversible. αMG mutase activity was assayed by coupling the formation of MeIT with the production of DMM in the presence of the isomerase. The purified mutase has an absolute requirement for cobamide coenzyme. The apparent Km values for αMG, α-(5,6-dimethylbenzimidazolyl)-cobamide coenzyme, α-(benzimidazolyl)-cobamide coenzyme, and α-(adenyl)-cobamide coenzyme are 7.14 x 10-3, 7.3 x 10-8, 3.0 x 10-7, and 1.25 x 10-6m, respectively. The most effective coenzyme, α-(5,6-dimethylbenzimidazolyl)cobamide coenzyme, is the same one that is synthesized in large amounts when C. barkeri is grown on nicotinic acid. No added cofactors are required for methylitaconate isomerase activity. The Km value is 6.2 x 10-3 m for methylitaconate and 4.0 x 10-3 m for dimethylmaleate. Both the mutase and the isomerase appear to be sulfhydryl proteins; neither requires metal ions for activity. A number of dicarboxylic acids are competitive inhibitors of αMG mutase but none of these affect MeIT isomerase activity.

Inhibition of cobamide coenzyme activity by a polypeptide derived from the cell wall of Lactobacillus leichmannii.

The cell wall and ribosomes of Lactobacillus leichmannii ATCC 7830, a vitamin B12-requiring organism, have been shown to inhibit the cobamide coenzyme-dependent propanediol hydro-lyase (EC 4.2.1.28) reaction. A B12-binder (polypeptide) of the wall was isolated and purified as a B12-polypeptide complex (4). The complex, when treated with an alkaline solution, resulted in detachment of B12 from the polypeptide. The free polypeptide, upon neutralization, could bind B12 again and cobamide coenzyme as well, thus capable of inhibiting the enzyme reaction. Comparison was also made of this polypeptide with a hog intrinsic factor preparation in the inhibition.

Effect of aglycone on the rate of cyanide cleavage of CoC-bond in nucleoside analogues of cobamide coenzyme.

Effect of aglycone on the rate of cyanide cleavage of CoC-bond in nucleoside analogues of cobamide coenzyme.

2,4-diaminovaleric acid: an intermediate in the anaerobic oxidation of ornithine by Clostridium sticklandii.

The oxidation of ornithine in the presence of proline by crude extracts of Clostridium sticklandii cells was stimulated by nicotinamide adenine dinucleotide, coenzyme A, alpha-ketoglutarate, dimethylbenzimidazolyl cobamide (DBC) coenzyme, MgCl(2), and adenosine diphosphate. Deletion of various cofactors resulted in the accumulation of a new dibasic amino acid which was identified as 2,4-diaminovaleric acid. Both the oxidation of ornithine to alanine and acetate and the conversion of ornithine to 2,4-diaminovaleric acid were stimulated by addition of DBC coenzyme, and both were inhibited by intrinsic factor, an inhibitor of cobamide coenzyme-dependent reactions. This inhibition was reversed by addition of DBC coenzyme. However, the oxidation of 2,4-diaminovaleric acid was insensitive to added intrinsic factor. The data indicate that 2,4-diaminovaleric acid represents the first intermediate in the oxidation of ornithine by C. sticklandii.

Ternary complex formation of 1,2-propanediol dehydratase, cobamide coenzyme and substrate analogue

Ternary complex formation of 1,2-propanediol dehydratase, cobamide coenzyme and substrate analogue

Spectrum and iron content of protein B2 from ribonucleoside diphosphate reductase.

Protein B2, one of the two non‐identical subunits of ribonucleoside diphosphate reductase from Escherichia coli B, contains two atoms of inorganic iron per molecule of protein. The metal could be removed from the protein either by prolonged dialysis against 8‐hydroxyquinoline or by precipitation with acid ammonium sulphate. This resulted in loss of enzyme activity which was recovered after treatment of the apoprotein with Fe2+ or Fe3+ but not with any other metals tested. Removal of iron did not change the behaviour of the apoprotein on sucrose gradient centrifugation or on polyacrylamide gel electrophoresis as compared to native protein B2.No “labile sulphide” could be demonstrated in protein B2.Protein B2 had a characteristic spectrum with a very sharp peak at 410 nm, a broader one around 360 nm and a steep shoulder at approximately 325 nm. The two peaks disappeared on removal of iron and reappeared on reactivation of the enzyme with the metal.When protein B2 was treated with hydroxyurea or hydroxylamine the peak at 410 nm (but not that at 360 nm) disappeared in parallel with enzyme activity. Such treatment did not result in the removal of iron.It is suggested that the very sharp peak at 410 nm originates from an unidentified component of protein B2. Furthermore, this component, together with iron, may be the functional equivalent in the E. coli system of cobamide coenzyme in ribonucleoside triphosphate reductase of Lacto‐bacillus leichmannii.

O-phosphorylethanolamine ammonia lyase, a new pyridoxal phosphate-dependent enzyme

O-phosphorylethanolamine ammonia lyase, a new enzyme present in vertebrate liver, has been shown to catalyze the catabolism of O-phosphorylethanolamine to equimolar quantities of ammonia, acetaldehyde, and inorganic phosphate. The enzyme occurs in the soluble fraction of the cell and has been partially purified from rabbit liver. Using 14C O-PE as substrate, the 14C acetaldehyde formed enzymatically was characterized as the 2,4-dinitrophenylhydrazone by infrared spectra and chromatography. Ethanolamine is not a substrate for the enzyme nor does the cobamide coenzyme affect its rate. The Km for O-phosphorylethanolamine was found to be 6.1 × 10 −4M and for pyridoxal phosphate 2.7 × 10 −7M.

The effect of cobamide coenzyme in methylmalonic acidemia.

Acta PaediatricaVolume 58, Issue 2 p. 178-180 THE EFFECT OF COBAMIDE COENZYME IN METHYLMALONIC ACIDEMIA B. LINDBLAD, B. LINDBLAD Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, Sweden Department of Physiological Chemistry Chemical Centre University of Lund Box 740 220 07 Lund 7 SwedenSearch for more papers by this authorK. LINDSTRAND, K. LINDSTRAND Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, SwedenSearch for more papers by this authorB. SVANBERG, B. SVANBERG Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, SwedenSearch for more papers by this authorR. ZETTERSTROM, R. ZETTERSTROM Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, SwedenSearch for more papers by this author B. LINDBLAD, B. LINDBLAD Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, Sweden Department of Physiological Chemistry Chemical Centre University of Lund Box 740 220 07 Lund 7 SwedenSearch for more papers by this authorK. LINDSTRAND, K. LINDSTRAND Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, SwedenSearch for more papers by this authorB. SVANBERG, B. SVANBERG Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, SwedenSearch for more papers by this authorR. ZETTERSTROM, R. ZETTERSTROM Departments of Physiological Chemistry, Chemical Centre, University of Lund, Lund and the Department of Pediatrics at Crown Princess Lovisa's Children's Hospital, Karolinska Institutet, Stockholm, SwedenSearch for more papers by this author First published: March 1969 https://doi.org/10.1111/j.1651-2227.1969.tb04704.xCitations: 28AboutPDF 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 Volume58, Issue2March 1969Pages 178-180 RelatedInformation

Deoxyribonucleotide Synthesis and the Role of Vitamin B12 in Erythropoiesis

Publisher Summary This chapter reviews the current knowledge of the several pathways of deoxyribonucleotide synthesis in bacteria and animal cells other than bone marrow and discusses the available evidence bearing on the pathway in human bone marrow. Current knowledge of the pathways of deoxyribonucleotide synthesis has been derived mainly from studies in two strains of bacteria—Escherichia coli, species lacking a nutritional requirement for vitamin B12 and Lactobacillus leichmannii, a nutritional vitamin B12 auxotroph. The system from E. coli utilizes ribonucleoside diphosphates as substrates; and L. leichmannii reduces ribonucleoside triphosphates. The more striking difference between the two enzyme systems is the fact that the enzyme from L. leichmannii, but not that from E. coli, has an absolute requirement for the vitamin B12 coenzyme, 5,6-dimethylbenzimidazolyl 5′-deoxyadenosine cobamide (DBCC). The ribonucleotide reductases from Novikoff hepatoma cells and E. coli have some properties in common that distinguish them from the corresponding enzyme of L. leichmannii, including (1) the preferred substrate is a ribonucleoside diphosphate, (2) no requirement for a cobamide coenzyme has been demonstrated, (3) an absolute requirement for Mg++, and (4) similar patterns of allosteric effects, among which is an inhibitory effect of deoxyadenosine triphosphate on the reductions of all four ribonucleotides.