1. Mobilization of copper ions from insoluble copper compounds by methanobactin Nay Myo Win, Joe Heimann, Shane Hodgson & Dr. Scott Hartsel  Chemistry Department  University of Wisconsin-Eau Claire Abstract Background Results MethodConclusion References  Copper is an essential nutrient for life as well as environmental toxin. Microbes are likely a very important part of the global copper cycle and the bioweathering of copper minerals. We have looked at the ability of a secreted bacterial copper-binding molecule, methanobactin, to mobilize copper from pure insoluble copper oxides and sulfides as models for bioweathering of corresponding natural copper minerals.  We followed the mobilization of copper spectrophotometrically in the presence of methanobactin and other known chemical copper binding agents. We find that methanobactin has a very high affinity for both Copper (I) and Copper(II) and can easily extract copper from all of these minerals, in particular Cu(I) minerals (saturation reached in less than 2 hours for Cu(I) minerals).  In addition unlike another Cu(I) chelating compound, BCS, methanobactin can stabilize the normally unstable Cu(I) ion for long periods of time even in the presence of water and oxygen.  These data suggest that in areas where these methanotrophic bacteria are common, secreted methanobactin could have a significant impact on soluble copper concentrations and weathering of copper- containing minerals.. Acknowledgements  University of Wisconsin – Eau Claire Chemistry Department  University of Wisconsin – Eau Claire Office of Research and Sponsored Programs (ORSP)  University of Wisconsin – Eau Claire Learning and Technology Services  What is methanobactin?  A polypeptide produced by the methanotrophs to collect copper from their surroundings  Can bind to other variety of earth metals with varying affinities  Why is it important?  Copper is necessary for methanotropic (methane- eating) bacteria to degrade methane and other organic molecules  Excess copper in the environment is toxic to living cells  Weathering processes  Oil spills cleaned up by methanotrophs  Extraction of copper from ores  Spectrophotometry The data were collected by using spectrophotometer. Different copper compounds, Copper (I) oxide, Copper (II) Oxide, Copper (I) sulfide, and Copper (II) sulfide, were used to determine the copper binding affinity of Bathocuproinedisulfonic acid (BCS), which is a known copper chelating agent, and methanobactin.  BCS - Copper bound BCS shows peak at 480 nm. - Spectra were taken 30 min/ cycle for 24 hours period.  Methanobactin - Copper bound mb loses absorbance at 394 nm. - Spectra were taken 15 min/ cycle for 24-48 hours period.  ICP-MS (Inductvely coupled plasma mass spectrometry) Copper compounds were incubated in MOPS buffer overnight and tested for availability of copper ions in water.  Methanobactin can rapidly remove copper ions from copper minerals and compounds normally considered to be completely insoluble in water.  The abundance of methanotrophs in nature suggests that MB could have a significant impact on mobilizing and weathering of copper minerals in the environment.  MB is at least as effective as the high-affinity specific Copper(I) chelator BCS at mobilizing copper and is more stable over time. Copper (I) Oxide Copper (II) Oxide Copper (I) Sulfide Copper (II) Sulfide Results from ICP-MS showing the insolubility of copper compounds in MOPS buffer Digenite (Cu9S5) with MB for 48 hours 1.Chi Fru, E. (2011). Copper Biogeochemistry: A Cornerstone in Aerobic Methanotrophic Bacterial Ecology and Activity. Geomicrobiology Journal, 28:601-614. 2.Campos, C., et. Al (2009). Evaluation of the copper (II) reduction assay using bathocuproinedisulfonic acid disodium salt for the total antioxidant capacity assessment: The CUPRAC-BCS assay. Analytical Biochemisty 392 (2009) 37- 44. Concentration (M)Standard Deviation (M) MOPS2.28E-083.93E-09 CuO3.30E-073.15E-09 Cu2O6.73E-062.60E-08 CuS9.87E-063.86E-08 Cu2S1.39E-054.56E-08