Laboratory investigations of copper acquisition by the aerobic methanotroph Methylosinus trichosporium OB3b and the ammonia oxidizing archaeon Nitrososphaera viennensis

Danielle D. Rushworth, Carolina Reyes, LEA STEINLE, Walter Schenkeveld, MORITZ F. LEHMANN, Christa Schleper, Stephan Krämer

Different microorganisms including archaea and bacteria require copper (Cu) as a metal co-factor for various metabolic pathways including aerobic methane oxidation and aerobic ammonia oxidation. Most known methanotrophs possess the Cu-bearing particulate methane monooxygenase (pMMO), an enzyme catalyzing the conversion of CH4 to methanol. Under Cu limitation, methanotrophic microorganisms can satisfy their relatively high Cu requirement through a high-affinity acquisition mechanism that involves the exudation of Cu-specific ligands, termed chalcophores.

Likewise, Cu is central to many enzymes in ammonia oxidizing archaea (AOA) including plastocyanins and multicopper oxidases. Although it is known that Cu plays an important role in AOA physiology and metabolism, it is not known how AOA acquire and transport Cu into the cell.

With respect to methanotrophs, we aim to elucidate the mechanisms involved in Cu acquisition from Cu-sulfide minerals using the aerobic CH4-oxidizing bacterium Methylosinus trichosporium OB3b. This will be investigated through kinetic dissolution and adsorption experiments conducted under oxic and anoxic conditions. Essential work has been done to isolate and purify methanobactin (mb) from the bacteria M.trichosporium following previously improved methods. A Cu(I)-sulfide phase, covellite, was synthesized for oxidative dissolution of Cu from the mineral. Results of oxidative dissolution will later be compared with the rates and mechanisms of ligand controlled dissolution of Cu from covellite using mb.

We have also explored the physiological response of the soil representative AOA isolate, Nitrososphaera viennensis (EN76)1,2 to Cu-limiting conditions. The chelator TETA (1,4,8,11-tetraazacyclotetradecane 1,4,8,11-tetraaceticacid hydrochloride hydrate) with selective affinity for Cu2+, was used to lower bioavailable Cu2+ in culture experiments as predicted by thermodynamic speciation calculations (PHREEQC). Ammonia and nitrite concentrations were measured in these cultures over time using colorimetric assays. Preliminary results show that N. viennensis is Cu-limited at free Cu2+concentrations as low as 6 x10-15 mol/L as compared to Cu-replete conditions (3 x 10-12 mol/L free Cu2+). This Cu2+ limiting threshold is higher than for denitrifying bacteria (<10-16 mol/L)4 and similar to other AOA1, which also possess Cu containing enzymes. Results from both studies will contribute to a better understanding of Cu acquisition by aerobic methanotrophs and archaea in environments where Cu availabilty is low.


Department für Ökogenomik und Systembiologie
Externe Organisation(en)
Universität Basel
ÖFOS 2012
Mikrobiologie, Umweltgeowissenschaften
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