Thallium-205 and carbon-13 NMR studies of human sero- and chicken ovotransferrin.

Biochemistry

Department of Biological Sciences, University of Calgary, Alberta, Canada.

Published: March 1994

We have examined the binding of Tl3+ to human serotransferrin and chicken ovotransferrin in the presence of carbonate and oxalate by 205Tl and 13C NMR spectroscopy. With carbonate as the synergistic anion, one observes two 205Tl NMR signals due to the bound metal ion in the two high-affinity iron-binding sites of each protein. When the same adducts are prepared with 13C-labeled carbonate, one finds two closely spaced doublets in the carbonyl region of the 13C NMR spectrum of serotransferrin; these correspond to the labeled anion directly bound to the metal ion in both sites of the protein. The analogous resonances in ovotransferrin are completely degenerate, and only one doublet can be detected. The magnitudes of the spin-spin coupling between the bound metal ion and carbonate range from 2J(205Tl-13C) approximately 270 to 290 Hz. We have used the proteolytic half-molecules of ovotransferrin and the recombinant N-terminal half-molecule of serotransferrin to assign the 205Tl and 13C NMR signals due to the bound metal ion and anion in both proteins. From titration studies, we found that Tl3+ is bound with a greater affinity at the C-terminal site of serotransferrin, whereas no site preference can be noted for ovotransferrin. When oxalate is used as the anion instead of carbonate, the 205Tl NMR signals arising from the bound metal ion in the sites of ovotransferrin are shifted downfield and become almost degenerate. A very complex pattern of resonances is observed for bound 13C2O4(2-) in the 13C NMR spectra of both proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

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http://dx.doi.org/10.1021/bi00177a022DOI Listing

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