Analysis of Cu(II) and Zn(II) binding of selected ambs as a function of varying pH and metal equivalents employing ion mobility mass spectrometric studies

Methanobactins (mb) are low molecular mass copper chelating chromopeptides called chalkophores secreted by methanotrophs. Methanotrophs utilize special enzymes known as methane mono-oxygenases for oxidation of methane. Methane mono-oxygenases are copper dependent and so, methanotrophs have developed the copper acquisition strategy by secreting mb's under lower copper conditions which can bind to copper(I) significantly. 2,3 mb originated by Methylosinus trichosporium (mb-OB3b) is the best chemically characterized methanobactin till date and is being used as prototype molecule. The mbs bind to variety of metals preferentially Cu(I) with high affinity. Our research group have designed a series of peptides based on the structure of mb-OB3b but have replaced the two enethiol-oxazolone groups with the alternative metal binding 2His-2Cys motif. My proposed research will involve studying Zn(II) binding affinity of analog methanobactin peptide-3 (amb3) peptide (acetyl-His1-Cys2-Gly3-Tyr4-Pro 5-His6-Cys7) as function of pH over a range of 3 to 11, and comparing it to the Zn(II) binding of amb1 (acetyl-His1 -Cys2-Gly3-Pro4-His5-Cys 6) and amb5 (acetyl-His1-Cys2-Gly3-Pro4-Tyr5-His6-Cys 7). The Cu(I/II) binding affinity and redox activity of the new alternative metal binding (amb6) peptide (acetyl-Cys1-His2-Cys 3-Gly4-Pro5-Cys6-His7-Cys 8) titrated with (0.2-1.00) molar equivalents of Cu(II) will also be studied employing ion mobility-mass spectrometry (IM-MS) techniques.