Journal
CANADIAN JOURNAL OF CHEMISTRY-REVUE CANADIENNE DE CHIMIE
Volume 89, Issue 7, Pages 779-788Publisher
CANADIAN SCIENCE PUBLISHING, NRC RESEARCH PRESS
DOI: 10.1139/V11-019
Keywords
quadrupolar central transition NMR; C-13 NMR; ovotransferrin; quadrupolar nuclei; metal ions; competition experiments
Categories
Funding
- Medical Research Council (MRC) of Canada
- AHFMR
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The transferrins are a family of relatively large bilobal proteins that play a major role in the transport of Fe3+, as well as several other physiological and nonphysiological metal ions. Transferrins can also act as antimicrobial agents, by tightly sequestering iron and making it unavailable for bacterial growth. Using a combination of quadrupolar central transition (QCT) metal ion NMR (Al-27, Sc-45, V-51, and Ga-71) and C-13 NMR, the binding and displacement of a variety of metal ions to ovotransferrin was studied through direct metal ion competition experiments. The metal ions investigated (Al3+, Co3+, Fe3+, Ga3+, In3+, Se3+, Y3+, and VO2+) were of differing ionic radius and charge, thus allowing for an assessment of how these factors contribute to metal ion affinity. The competition for the N- and C-terminal metal ion binding sites on ovotransferrin was directly followed by metal ion QCT NMR. Moreover, C-13 NMR was used to study the two protein-bound synergistic anions (C-13-labeled carbonate), whose chemical shifts are distinct and dependent on the bound metal ion that is present in the binding sites. The observed order of decreasing affinity for the metal ions studied was Fe3+ approximate to In3+ >= Sc3+ >= Ga3+ > Al3+ > VO2+ > Y3+ >= Co3+. These results illustrate how a combination of multinuclear solution NMR methods can provide unique insights into the ligand binding properties of larger metalloproteins.
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