The binding of several corrinoids to the binding site of human intrinsic factor, transcobalamin or haptocorrin was investigated. p-Cresolyl cobamide and 2-amino-vitamin B1 are complete corrinoids, whose nucleotide at the lower face of the corrin ring is not coordinated to the cobalt. These corrinoids were 2 lo3 times less efficiently recognized by intrinsic factor or transcobalamin than vitamin BIZ, which contains a Co-coordinated nucleotide. Pseudovitamin B12, with a weak Co-N coordination bond, revealed only moderate affinity to intrinsic factor. From these findings it is concluded that the cobamide binding to intrinsic factor and transcobalamin is strongly affected by the Co-N coordination bonds of their lower cobalt nucleotide ligands. We suggest that the Co-N coordination bond positions the nucleotide at a critical distance to the corrin ring, which is recognized by the binding proteins.Human haptocorrin, however, disclosed to distinctive selectivity regarding the different corrinoid structures. The protein bound all corrinoids with similar efficiency, independent of the strength of their Co-N coordinations, or the structures of their lower Cocl ligands. Hence, the corrin ring, rather than a structural feature induced by the Co-N coordination, has to be considered responsible for the corrinoid binding to haptocorrin.Cobalamin-dependent methionine synthase, methylmalonyl-CoA mutase and leucine 2,3-aminomutase reactions in eukaryotic cells require a sufficient vitamin B12 supply by corrinoid-binding and corrinoid-transporting proteins. Intrinsic factors, transcobalamins and haptocorrins meet these needs in humans and other mammalia [I]. Homogeneous preparations of the human corrinoid binding proteins were obtained from gastric juice and serum [I]. Yet, the cobamide binding mechanism of these proteins remained equivocal [l-41, as well as the chemical structure, the source and the function of corrinoid analogues in human serum [5] and animal tissues [3, 61.Two features of the vitamin B I 2 molecule could in principle provide the structural prerequisites for the corrinoid binding by intrinsic factor, transcobalamin and haptocorrin: the corrin ring and the upper or lower cobalt ligands.From previous cobamide binding studies with benzimidazolyl cobamide, 5-hydroxybenzimidazolyl cobamide and 5-methoxybenzimidazolyl cobamide it was considered that an unmodified nucleotide group with 5,6-dimethylbenzimidazole as the heterocyclic base below the corrin ring is sufficient for cobalamin binding to intrinsic factor [7]. According to Grasbeck's corrinoid binding model, the upper cobalt ligandCorrespondence to E. Stupperich, Angewandte Mikrobiologie, Universitat Ulm, Albert-Einstein-Allee 11, W-7900 Ulm, Federal Republic of Germany Abbreviations. hIF, human intrinsic factor; hHC, human heptocorrin; hTC, human transcobalamin; FAB-MS, fast-atom-bombardment mass spectroscopy.Enzymes. Methionine synthase (EC 2.1.1.13); methylmdlonylCoA mutase (EC 5.4.99.2); leucine 2,3-aminomutase (EC 5.4.3.7).(e. g. a cyano, hydroxyl or adenos...
Growing cells of Propionibacterium shermanii, when supplemented with imidazole, produced a "complete" corrinoid, as reported from a similar (but also cobinamide supplemented) setup by Müller and co-workers (Eberhard, G.; et al. Biol. Chem. Hoppe-Seyler 1988, 369,1091. The corrinoid isolate from workup with added potassium cyanide was confirmed to consist mainly (84%) of the suggested Co/3-cyanoimidazolylcobamide (1), the vitamin Bn analogue, in which the cobalt-coordinating 5,6-dimethylbenzimidazole base of the vitamin is replaced by imidazole. An extensive NMR spectroscopic analysis of aqueous solutions of 1 was carried out: all and 13C signals in the NMR spectra of 1 were assigned unambiguously, exploiting two-dimensional gradient enhanced homonuclear and heteronuclear experiments, including a new variant of the two-dimensional H-relayed , -Overhauser experiment at natural abundance, called 2D-HSQC-ROESY, which combines the merits of heteronuclear single quantum coherence (HSQC) and rotating frame Overhauser spectroscopy (ROESY). The structural analyses by NMR, UV, and CD spectroscopy of 1 in aqueous solution did not unravel any gross structural differences of the corrin ligand and of the nucleotide loop of 1 and of vitamin B]2 (2). Cryo-temperature single-crystal structure analyses have been carried out for 1 and 2, the latter one to obtain reference data more accurate than the ones available from Hodgkin's analysis (Brink-Shoemaker, C. et al. Proc. R. Soc. London, A 1964,278,1). The imidazolylcobamide 1 crystallized at room temperature from aqueous acetone in the orthorhombic space group P2{2{2\, with Z = 4. Diffraction data were collected at 92 K (a = 15.335( 16) Á, b = 21.974(9) Á, c = 25.501(11) Á, V = 8705(2) Á3), the structure was solved by the Patterson method and refined to a final R value of 0.1017 for 6105 reflections with F0 > 4
The Cop-cyanocobamides obtained by cyanide extractions from several acetogenic bacteria were structurally characterized by ultraviolet/visible spectra, proton-nuclear-magnetic-resonance spectra and fast-atom-bombardment mass spectra. p-Cresolylcobamide was detected as a major corrinoid from Sporomusa ovata. This 'complete' corrinoid was isolated from an organism for the first time. Instead of the common Coa bases of the known and biologically active cobamides, p-cresolylcobamide contained a glycosidically bound cresolyl function that was unable to coordinate to the cobalt of the corrin ring. An additional, previously unknown corrinoid from natural sources, Coa-[a-(5-methoxy-6-methylbenzimidazolyl)]-Co~-cyanocobamide, was isolated along with vitamin B1 from Clostridium formicoaceticum. Both homoacetogenic eubacteria were grown on methanol and contained high amounts of corrinoids (> 950 nmol/g cell dry mass). Less corrinoid was isolated from Acetobacterium woodii and characterized as vitamin B12.
Cultures of the autotrophic bacterium Methanobacterium thermoautotrophicum were shown to assimilate acetate when grown on CO2 and H2 in the presence of acetate. At 1 mM acetate 10% of the cell carbon came from acetate, the rest from CO2. At higher concentrations the percentage increased to reach a maximum of 65% at acetate concentrations higher than 20 mM. The data suggest that acetate may be an important carbon source under physiological conditions. The incorporation of acetate into alanine, aspartate and glutamate was studied in more detail. The cells were grown on CO2 and H2 in the presence of 1 mM U-14C-acetate. The three amino acids were isolated from the labelled cells by a simplified procedure. Alanine, aspartate and glutamate were found to have the same specific radioactivity. Degradation studies showed that C1 of alanine, C1 and C4 of aspartate, and C1 and C5 of glutamate were exclusively derived from CO2, whereas C2 and C3 of alanine and aspartate, and C3 and C4 of glutamate were partially derived from acetate. These findings and the presence of pyruvate synthase, phosphoenolpyruvate carboxylase and alpha-ketoglutarate synthase in M. thermoautotrophicum indicate that CO2 is assimilated into the three amino acids via acetyl CoA carboxylation to pyruvate, phosphoenolpyruvate carboxylation to oxaloacetate, and succinyl CoA carboxylation to alpha-ketoglutarate.
Cobalamin and the native and diepimeric forms of factor F430 catalyzed the reductive dechlorination of 1,2-dichloroethane (1,2-DCA) to ethylene or chloroethane (CA) in a buffer with Ti(III) citrate as the electron donor. Ethylene was the major product in the cobalamin-catalyzed transformation, and the ratio of ethylene to CA formed was 25:1. Native F430 and 12,13-di-epi-F430 produced ethylene and CA in ratios of about 2:1 and 1:1, respectively. Cobalamin dechlorinated 1,2-DCA much faster than did factor F430. Dechlorination rates by all three catalysts showed a distinct pH dependence, correlated in a linear manner with the catalyst concentration and doubled with a temperature increase of 10°C. Crude and boiled cell extracts of Methanosarcina barkeri also dechlorinated 1,2-DCA to ethylene and CA with Ti(III) citrate as the reductant. The catalytic components in boiled extracts were heat and oxygen stable and had low molecular masses. Fractionation of boiled extracts by a hydrophobic interaction column revealed that part of the dechlorinating components had a hydrophilic and part had a hydrophobic character. These chemical properties of the dechlorinating components and spectral analysis of boiled extracts indicated that corrinoids or factor F430 was responsible for the dechlorinations. The ratios of 3:1 to 7:1 of ethylene and CA formed by cell extracts suggested that both cofactors were concomitantly active.Chlorinated aliphatic C1 and C2 hydrocarbons, widespread contaminants in different environments, are found to be reductively dechlorinated by pure cultures of methanogens, sulfate reducers, homoacetogens, and other anaerobic bacteria (4,11,13,16,18,22,26,33). Tetrachloromethane (CCd4) was found to be transformed to lower-chlorinated methanes and CO2 by native or autoclaved cell suspensions of methanogens, Acetobacterium woodii, and Desulfobacterium autotrophicum (12,13,26). These anaerobic bacteria fix CO2 via the acetyl-coenzyme A pathway (Wood pathway) or degrade acetate via a reversed acetyl-coenzyme A pathway (17,32,42,44). Carbon monoxide dehydrogenase and a corrinoid-iron-sulfur protein are central enzymes of this pathway. It has been suggested that the anaerobic transformation of CCl4 could be a cometabolic activity of Wood pathway enzymes (12, 13). Free cobalamin catalyzed reductive dechlorination of CCl4 in a buffer reduced with Ti(III) citrate as the electron donor (27). In addition to lowerchlorinated methanes, cobalamin produced CO from CCl4 (28), a compound which is oxidized to CO2 by carbon monoxide dehydrogenase (17,32,42,44). In methanogenic bacteria, factor F430 may also be involved in reductive dechlorination. Factor F430 is a cofactor of methyl-coenzyme M reductase, the enzyme which catalyzes the last step in methanogenesis from methyl-coenzyme M to CH4 (14). This cofactor reductively dechlorinated CCl4 to lower-chlorinated methanes (26). All Cell suspensions of methanogens reductively dechlorinated 1,2-dichloroethane (1,2-DCA) to ethylene and chloroethane (CA) (22). We demonstrate here th...
The cmuA and cmuB genes are required for growth of Methylobacterium chloromethanicum strain CM4 with chloromethane as the sole carbon source. While CmuB was previously shown to possess methylcobalamin:tetrahydrofolate methyltransferase activity, sequence analysis indicated that CmuA represented a novel and so far unique two-domain methyltransferase/corrinoid-binding protein involved in methyl transfer from chloromethane to a corrin moiety. CmuA was purified from wild-type M. chloromethanicum strain CM4 and characterized as a monomeric, cobalt-containing and zinc-containing enzyme of molecular mass 67 kDa with a bound vitamin B 12 cofactor. In combination, CmuA and CmuB proteins catalyze the in vitro transfer of the methyl group of chloromethane to tetrahydrofolate, thus affording a direct link between chloromethane dehalogenation and core C1 metabolism of Methylobacterium. Chloromethane dehalogenase activity in vitro is limited by CmuB, as formation of methyltetrahydrofolate from chloromethane displays apparent Michaelis±Menten kinetics with respect to methylated CmuA, with an apparent K m of 0.27 mm and a V max of 0.45 U´mg 21 . This contrasts with sequence-related systems for methyl transfer from methanogens, which involve methyltransferase and corrinoid protein components in well-defined stoichiometric ratios.
The corrinoids from the obligate anaerobe Clostridium cochlearium were extracted as a mixture of Co  -cyano derivatives. From 50 g of frozen cells, approximately 2 mg (1.5 mol) of B 12 derivatives was obtained as a crystalline sample. Analysis of the corrinoid sample of C. cochlearium by a combination of high-pressure liquid chromatography and UV-Vis absorbance spectroscopy revealed the presence of three cyano corrinoids in a ratio of about 3:1:1. The spectroscopic data acquired for the sample indicated the main components to be pseudovitamin B 12 (Co  -cyano-7؆-adeninylcobamide) (60%) and factor A (Co  -cyano-7؆-[2-methyl]adeninylcobamide) (20%). Authentic pseudovitamin B 12 was prepared by guided biosynthesis from cobinamide and adenine. Both pseudovitamin B 12 and its homologue, factor A, were subjected to complete spectroscopic analysis by UV-Vis, circular dichroism, mass spectrometry, and by one-and two-dimensional 1 H, 13 C-, and 15 N nuclear magnetic resonance (NMR) spectroscopy. The third component was indicated by the mass spectra to be an isomer of factor A and is likely (according to NMR) to be 7؆-[N 6 -methyl]-adeninylcobamide, a previously unknown corrinoid. C. cochlearium thus biosynthesizes as its native "complete" B 12 cofactors the 7؆-adeninylcobamides and two homologous corrinoids, in which the nucleotide base is a methylated adenine.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.