Frederick S. Archibald

Manganese poisoning and the attack of trivalent manganese upon catecholamines

Human manganese poisoning or manganism results in damage to the substantia nigra of the brain stem, a drop in the level of the inhibitory neurotransmitter dopamine, and symptoms resembling those of Parkinsons disease. Manganic (Mn3+) manganese ions were shown to be readily produced by O-2 in vitro and spontaneously under conditions obtainable in the human brain. Mn3+ as its pyrophosphate complex was shown to rapidly and efficiently carry out four-electron oxidations of dopamine, its precursor dopa (3,4-dihydroxyphenylalanine), and its biosynthetic products epinephrine and norepinephrine. Mn3+-pyrophosphate was shown to specifically attack dihydroxybenzene derivatives, but only those with adjacent hydroxyl groups. Further, the addition of Mn2+-pyrophosphate to a system containing a flux of O2- and dopamine greatly accelerated the oxidation of dopamine. The oxidation of dopamine by Mn3+ neither produced nor required O2, and Mn3+ was far more efficient than Mn2+, Mn4+ (MnO2), O2-, or H2O2 in oxidizing the catecholamines. A higher oxidation state, Mn(OH)3, formed spontaneously in an aqueous Mn(OH)2 precipitate and slowly darkened, presumably being oxidized to MnO2. Like reagent MnO2, it weakly catalyzed dopamine oxidation. However, both MnO2 preparations showed dramatically increased abilities to oxidize dopamine in the presence of pyrophosphate due to enhancement of the spontaneous formation of the Mn3+ complex. These results strongly suggest that the pathology of manganese neurotoxicity is dependent on the ease with which simple Mn3+ complexes are formed under physiological conditions and the efficiency with which they destroy catecholamines.

Manganese: Its Acquisition by and Function in the Lactic Acid Bacteria

The transition metal manganese is considered to be a minor micronutrient in both pro- and eukaryotes, usually being required from the environment at subnanomolar levels. Until recently, Mn was only known to function in cells as a cofactor for a few enzymatic reactions. A notable exception has been reported in many lactic acid bacterial species which require micromolar medium Mn levels for growth and contain up to 35 mM Mn. These high Mn concentrations are accompanied by the near or complete absence of intracellular iron and superoxide dismutase (SOD). Lacking hemes, Lactobacillus plantarum and related species contain a unique Mn-cofactored catalase as well as millimolar Mn(II) in a nonenzymic complex performing the function of the micromolar superoxide dismutase found in most other aerotolerant cells. The high Mn(II) levels are accumulated via an efficient active transport system and are stored intracellularly in a high molecular weight complex. Study of Lactobacillus plantarum has provided an interesting example of the substitution of Mn for Fe in several of the biological roles of Fe, an alternative mechanism of aerotolerance, and a better understanding of the unique biochemistry of the lactic acid bacteria.

Isolation of Poly-3-Hydroxybutyrate Metabolism Genes from Complex Microbial Communities by Phenotypic Complementation of Bacterial Mutants

ABSTRACT The goal of this study was to initiate investigation of the genetics of bacterial poly-3-hydroxybutyrate (PHB) metabolism at the community level. We constructed metagenome libraries from activated sludge and soil microbial communities in the broad-host-range IncP cosmid pRK7813. Several unique clones were isolated from these libraries by functional heterologous complementation of a Sinorhizobium meliloti bdhA mutant, which is unable to grow on the PHB cycle intermediate d-3-hydroxybutyrate due to absence of the enzyme d-3-hydroxybutyrate dehydrogenase activity. Clones that conferred d-3-hydroxybutyrate utilization on Escherichia coli were also isolated. Although many of the S. meliloti bdhA mutant complementing clones restored d-3-hydroxybutyrate dehydrogenase activity to the mutant host, for some of the clones this activity was not detectable. This was also the case for almost all of the clones isolated in the E. coli selection. Further analysis was carried out on clones isolated in the S. meliloti complementation. Transposon mutagenesis to locate the complementing genes, followed by DNA sequence analysis of three of the genes, revealed coding sequences that were broadly divergent but lay within the diversity of known short-chain dehydrogenase/reductase encoding genes. In some cases, the amino acid sequence identity between pairs of deduced BdhA proteins was <35%, a level at which detection by nucleic acid hybridization based methods would probably not be successful.

Coliform Bacteria and Nitrogen Fixation in Pulp and Paper Mill Effluent Treatment Systems

ABSTRACT The majority of pulp and paper mills now biotreat their combined effluents using activated sludge. On the assumption that their wood-based effluents have negligible fixed N, and that activated-sludge microorganisms will not fix significant N, these mills routinely spend large amounts adding ammonia or urea to their aeration tanks (bioreactors) to permit normal biomass growth. N2 fixation in seven Eastern Canadian pulp and paper mill effluent treatment systems was analyzed using acetylene reduction assays, quantitative nitrogenase (nifH) gene probing, and bacterial isolations. In situ N2 fixation was undetectable in all seven bioreactors but was present in six associated primary clarifiers. One primary clarifier was studied in greater detail. Approximately 50% of all culturable cells in the clarifier contained nifH, of which >90% were Klebsiella strains. All primary-clarifier coliform bacteria growing on MacConkey agar were identified as klebsiellas, and all those probed contained nifH. In contrast, analysis of 48 random coliform isolates from other mill water system locations showed that only 24 (50%) possessed thenifH gene, and only 13 (27%) showed inducible N2-fixing activity. Thus, all the pulp and paper mill primary clarifiers tested appeared to be sites of active N2fixation (0.87 to 4.90 mg of N liter−1 day−1) and a microbial community strongly biased toward this activity. This may also explain why coliform bacteria, especially klebsiellas, are indigenous in pulp and paper mill water systems.