Methanotrophic bacteria catalyze the aerobic oxidation of methane to methanol using enzymes with copper (Cu)-based active sites. To facilitate the acquisition of Cu ions some methanotrophic bacteria secrete small postranslationally modified peptides known as methanobactins. Analogous to siderophores and iron, methanobactin strongly binds Cu and functions as an extracellular Cu recruitment relay. In addition to binding copper, methanobactins will bind most transition metals and near-transition metals and protect the host methanotroph as well as other bacteria from metal toxicity. Recent studies have demonstrated that some methanotrophs are capable of degrading MeHg in aerobic environments. The metal binding site is comprised of two enethiolate groups each conjugated with nitrogen-containing heterocycles. The unique chemistry of these functional groups allows methanobactins to bind a wide range of transition metal ions. In addition, the complexation of metal ions is reflected in the electronic structure of the conjugated system, thus absorbance and fluorescence spectra of methanobactins are sensitive to the coordination state of the peptide. We report the thermodynamics of transition metal ion complexation by methanobactin from <i>Methylocystis sp.</i> strain SB2 using isothermal titration calorimetry (ITC). Complexation of methanobactin with transition metal ions entails a range of intra- and intermolecular interactions depending on the metal:methanobactin ratio. We determine thermodynamic parameters such as enthalpy, entropy, stoichiometry, and binding constants for several group 11 and 12 transition metals and compare the results to data obtained from spectroscopic investigations. Absorbance, fluorescence, extended X-ray absorption fine structure (EXAFS) spectroscopy and TD-DFT calculations provide insights into coordination numbers and geometries of the metal binding site. In addition, we characterize the competitive interactions between complexes of methanobactin with transition metal ions and model compounds representing several classes of functional groups, such as thiols, thiones, imines, amines, and the widely used chelating ligand ethylene diamine tetraacetic acid (EDTA). The present data offer insights into mechanistic aspects of transition metal complexation by the chalkophore methanobactin and suggest that it may influence the speciation and biogeochemical cycling of several transition metals in addition to Cu.

Mechanistic insights into the interactions of methanobactins with group 11 and 12 transition metals

ACS Fall 2021

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