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Citrobacter amalonaticus Y19 (Y19) was isolated because of its ability for carbon monoxide-dependent hydrogen production (water-gas shift reaction). This paper reports the assimilation of glycerol and the production of 1,3-propanediol (1,3-PDO) by Y19. Genome sequencing revealed that Y19 contained the genes for the utilization of glycerol and 1,2-propanediol ( pdu operon) along with those for the synthesis of coenzyme B ( cob operon). On the other hand, it did not possess the genes for the fermentative metabolism of glycerol of Klebsiella pneumoniae, which consists of both the oxidative ( dhaD and dhaK) and reductive ( dhaB and dhaT) pathways. In shake-flask cultivation under aerobic conditions, Y19 could grow well with glycerol as the sole carbon source and produced 1,3-PDO. The level of 1,3-PDO production was improved when vitamin B was added to the culture medium under aerobic conditions. Under anaerobic conditions, cell growth and 1,3-PDO production on glycerol was also possible, but only when an exogenous electron acceptor, such as nitrate or fumarate, was added. This is the first report of the glycerol metabolism and 1,3-PDO production by C. amalonaticus Y19. [ABSTRACT FROM AUTHOR]

Computational modeling of the enzymatic activity of B12-dependent enzymes requires a detailed understanding of the factors that influence the strength of the Co&bond;C bond and the limits associated with a particular level of theory. To address this issue, a systematic analysis of the electronic and structural properties of coenzyme B12 models has been performed to establish the performance of three different functionals including B3LYP, BP86, and revPBE. In particular the cobalt–carbon bond dissociation energies, axial bond lengths, and selected stretching frequencies have been analyzed in detail. Current analysis shows that widely used B3LYP functional significantly underestimates the strength of the Co&bond;C bond while the nonhybrid BP86 functional produces very consistent results in comparison to experimental data. To explain such different performance of these functionals molecular orbital analysis associated with axial bonds has been performed to show differences in axial bonding provided by hybrid and nonhybrid functionals. © 2006 Wiley Periodicals, Inc. J Comput Chem 27: 1429–1437, 2006 [ABSTRACT FROM AUTHOR]

Abstract: Heterodisulfide reductase (HDR) catalyzes the formation of coenzyme M (CoM–SH) and coenzyme B (CoB–SH) by the reversible reduction of the heterodisulfide, CoM–S–S–CoB. This reaction recycles the two thiol coenzymes involved in the final step of microbial methanogenesis. Electron paramagnetic resonance (EPR) and variable-temperature magnetic circular dichroism spectroscopic experiments on oxidized HDR incubated with CoM–SH revealed a S =1/2 [4Fe–4S]3+ cluster, the EPR spectrum of which is broadened in the presence of CoM–33SH [Duin, E.C., Madadi-Kahkesh, S., Hedderich, R., Clay, M.D. and Johnson, M.K. (2002) Heterodisulfide reductase from Methanothermobacter marburgensis contains an active-site [4Fe–4S] cluster that is directly involved in mediating heterodisulfide reduction. FEBS Lett. 512, 263–268; Duin, E.C., Bauer, C., Jaun, B. and Hedderich, R. (2003) Coenzyme M binds to a [4Fe–4S] cluster in the active site of heterodisulfide reductase as deduced from EPR studies with the [33S]coenzyme M-treated enzyme. FEBS Lett. 538, 81–84]. These results provide indirect evidence that the disulfide binds to the iron–sulfur cluster during reduction. We report here direct structural evidence for this interaction from Se X-ray absorption spectroscopic investigation of HDR treated with the selenium analog of coenzyme M (CoM–SeH). Se K edge extended X-ray absorption fine structure confirms a direct interaction of the Se in CoM–SeH-treated HDR with an Fe atom of the Fe–S cluster at an Fe–Se distance of 2.4Å. [Copyright &y& Elsevier]

Recombinant glycerol dehydratase of Klebsiella pneumoniae was purified to homogeneity. The subunit composition of the enzyme was most probably α2 β2 γ2 . When (R )- and (S )-propane-1,2-diols were used independently as substrates, the rate with the (R )-enantiomer was 2.5 times faster than that with the (S )-isomer. In contrast to diol dehydratase, an isofunctional enzyme, the affinity of the enzyme for the (S )-isomer was essentially the same or only slightly higher than that for the (R )-isomer (K m(R ) /K m(S ) = 1.5). The crystal structure of glycerol dehydratase in complex with cyanocobalamin and propane-1,2-diol was determined at 2.1 Å resolution. The enzyme exists as a dimer of the αβγ heterotrimer. Cobalamin is bound at the interface between the α and β subunits in the so-called ‘base-on’ mode with 5,6-dimethylbenzimidazole of the nucleotide moiety coordinating to the cobalt atom. The electron density of the cyano group was almost unobservable, suggesting that the cyanocobalamin was reduced to cob(II)alamin by X-ray irradiation. The active site is in a (β/α)8 barrel that was formed by a central region of the α subunit. The substrate propane-1,2-diol and essential cofactor K+ are bound inside the (β/α)8 barrel above the corrin ring of cobalamin. K+ is hepta-coordinated by the two hydroxyls of the substrate and five oxygen atoms from the active-site residues. These structural features are quite similar to those of diol dehydratase. A closer contact between the α and β subunits in glycerol dehydratase may be reminiscent of the higher affinity of the enzyme for adenosylcobalamin than that of diol dehydratase. Although racemic propane-1,2-diol was used for crystallization, the substrate bound to glycerol dehydratase was assigned to the (R )-isomer. This is in clear... [ABSTRACT FROM AUTHOR]

Cells of Synechococcus sp., a rod-shaped, unicellular cyanobacterium (blue-green alga) can be readily snychronized by depriving the cells of carbon dioxide and light for a 12 h period. On resumption of growth, a portion of the population undergoes two sharply synchronized divisions. Ribonucleotide reductase activity was found to be maximal during the time of DNA synthesis in these cells. The peak of reductase activity could be abolished by adding inhibitors such a chloramphenicol to the culture, suggesting that the enzyme is induced at the gene level in the cyanobacteria. Additional properties of ribonucleotide reductase were investigated in Synechococcus cells made permeable by treatment with ether. Cytidine triphosphate reduction is absolutely dependent on adenosylcobalamin (coenzyme B) and is subject to allosteric stimulation by deoxyadenosine triphosphate. [ABSTRACT FROM AUTHOR]