Spectral and copper binding properties of methanobactin from the facultative methanotroph Methylocystis strain SB2

Methanobactin (mb) is the first characterized example of a chalkophore, a class of copper-binding chromopeptides similar to iron-binding siderophores. Structural, redox, themodynamic, and spectral studies on chalkophores have focused almost exclusively on the mb from Methylosinus trichosporium OB3b (mb-OB3b). The structural characterization of a second mb from Methylocystis strain SB2 (mb-SB2) provides a means to examine the core structural features and metal binding properties of this group of chromopeptides. With the exception of the 5-membered rings (either oxazolone or imidazolone), enethiol groups, and the N-terminus oxo group, the structure of mb-SB2 differs markedly from mb-OB3b. In particular the amino acids commonly associated with metal coordination and redox activity found in mb-OB3b, Cys, Met, and Try, are replaced by Ala or are missing in mb-SB2. In this report the spectral and thermodynamic properties of mb-SB2 are presented and compared to mb-OB3b. The results demonstrate that the spectral and basic copper binding properties of both methanobactins are similar and the unique copper binding capacity of both methanobactins lies primarily in the pair of five-membered rings and associated enethiol groups. The remaining portions of the methanobactin appear to provide the scaffolding that brings together of the two ring systems to produce the tetrahedral binding site for copper binding.

Top: Core features of methanobactin from Methylocystis strain SB2 and Methylosinus trichosporium OB3b. Bottom left: UV–visible absorption spectra of methanobactin from Methylocystis strain SB2. Bottom right: Fluorescence spectra of methanobactin from Methylocystis strain SB2.Figure optionsDownload as PowerPoint slideHighlights
► Copper binding capacity of mb-SB2 is similar but not identical to mb-OB3b.
► The unique copper binding capacity of mbs resides in the rings and enethiol groups.
► Displacement ITC is a viable method to measure binding constants > 1012.