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Trace element concentrations vary substantially during snowmelt runoff, with changes in the dissolved versus particulate fractions potentially impacting their movement at the catchment scale. To investigate trace element behavior in a snowmelt dominated stream, we measured concentrations in different size fractions in the Provo River (northern Utah, USA). We sampled the river at three locations during water years 2016-2018 and 2021-2023 for trace metal and major ion concentrations. During the final year, we collected three fractions (unfiltered, <0.45 µm filtered, and <0.22 µm filtered) for trace metal chemistry and calculated the particulate concentration as the difference between the unfiltered and 0.45 µm fraction. Stream measurements included pH, discharge, turbidity, and fluorescent dissolved organic matter (fDOM). We also sampled water sources (soil water, ephemeral streams, and snowpack) to compare trace metal concentrations across the watershed with the stream. Trace metal (Al, Be, Fe, Pb) and rare earth element (REE +Y) concentrations had the highest particulate and dissolved concentrations during snowmelt runoff. In contrast, major cations were primarily found in the dissolved fraction with lowest concentrations during snowmelt runoff. Major cation and particulate metal concentrations increased from upstream to downstream. The increased trace metal and REE + Y concentrations during snowmelt relative to baseflow may be explained by increased discharge and associated increased turbidity and fDOM. A comparison with water sources suggests that dissolved and particulate trace metals in the stream are sourced from flushed soil water, as trace metal transport through the watershed is facilitated by suspended sediment and dissolved organic matter. For most of the trace metals and REE +Y, concentrations were similar in the <0.45 µm and <0.22 µm fractions. Yet Al tended to have higher concentrations in the <0.45 µm fraction, suggesting a colloidal form of Al between 0.22 µm and 0.45 µm may exist. Differences from upstream to downstream may be related to changes in pH, from ~7 at the upper site to ~8 at the lower sites, which would change sorption capacity, saturation indices, or speciation. Our study demonstrates that trace metal concentrations are variable during snowmelt as the metals interact with suspended particles or dissolved organic matter and are influenced by changing water chemistry, with implications for understanding water quality impairments in snowmelt-dominated streams.

CopA, a thermophilic membrane ATPase from Archaeoglobus fulgidus, drives the outward movement of Cu+ across cellular membranes. CopA contains at least two metal binding domains, a regulatory N-terminal Metal Binding Domain (N-MBD) and an occlusion/coordinating metal binding site in the 6th, 7th and 8th transmembrane segments. Previous studies showed that the presence of millimolar concentration of Cys is essential for CopA activity. The high affinity of CopA for metal in the presence of millimolar concentration of Cys suggests a multifaceted interaction of the enzyme with Cys. To elucidate the role of Cys, we studied its effect on the partial reactions of the catalytic cycle of CopA. We observed that 2-50 mM Cys accelerates enzyme turnover with little effect on the Cu+ affinity of CopA. Cys accelerates enzyme phosphorylation, but has no effect on the dephosphorylation rates. Thus, Cys increases steady state phosphoenzyme levels. Besides, Cys has no significant effect on E1¡ÃƒÂªE2 equilibrium. Similar results were observed in truncated CopA lacking the N-MBD suggesting that enzyme activation by Cys is independent of the regulatory metal binding sites. These results and the kinetic analysis of activation curves suggest that while Cu+ is delivered to the transport site as a Cu-Cys complex, Cys in the mM range stimulates the ATPase acting as a non-essential activator.

Many processes in living systems occur through transient interactions among proteins. Those interactions are often weak and are driven by small changes in free energy. Due to the short-living nature of these interactions, our knowledge about driving forces, dynamics and structures of these types of protein-protein heterocomplexes are though limited. This is especially important for cellular copper (Cu) trafficking: Copper ions are essential for all eukaryotes and most bacteria. As a cofactor in many enzymes, copper is especially vital in respiration or detoxification. Since the same features that make copper useful also make it toxic, it needs to be controlled tightly. Additionally, in the reducing environment of the cytosol, Cu is present as insoluble Cu(I). To circumvent both toxicity and solubility issues, a system has evolved where copper is comforted by certain copper binding proteins, so-called Cu-chaperones. They transiently interact with each other to distribute the Cu atoms in a cell. In humans, one of them is Atox1. It binds copper with a binding site containing two thiol residues and transfers it to other binding sites, mostly those of a copper pump, ATP7B (also known as Wilsons disease protein). My work was aimed at understanding copper-mediated protein-protein interactions on a molecular and mechanistic level. Which amino acids interact with the metal? Which forces drive the transfer from one protein to the other? Using biophysical and biochemical methods such as chromatography and calorimetry on wild type and point-mutated proteins in vitro, we found that the copper is transferred via a dynamic intermediate complex that keeps the system flexible while shielding the copper against other interactions. Although similar transfer interactions can be observed in other organisms, and many conclusions in the copper field are drawn from bacterial and yeast analogs, we believe that it is important to investigate human proteins, too. Not only is their regulation different, but also only in humans we find the diseases linked to the proteins: Copper level regulation diseases are to be named first, but atypical copper levels have also been linked to tumors and amyloid dispositions. In summary, my observations and conclusions are of basic research character and can be of importance for both general copper and human medicinal research.

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