Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 100, Issue 7, Pages 3659-3664Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0536897100
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- NIDDK NIH HHS [R01 DK15056] Funding Source: Medline
- PHS HHS [R29 A132226] Funding Source: Medline
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Although the presence of an exogenous anion is a requirement for tight Fe3+ binding by the bacterial (Neisseria) transferrin nFbp, the identity of the exogenous anion is not specific in vitro. nFbp was reconstituted as a stable iron containing protein by using a number of different exogenous anions [arsenate, citrate, nitrilotriacetate, pyrophosphate, and oxalate (symbolized by X)] in addition to phosphate, predominantly present in the recombinant form of the protein. Spectroscopic characterization of the Fe3+/anion interaction in the reconstituted protein was accomplished by UV-visible and EPR spectroscopies. The affinity of the protein for Fe3+ is anion dependent, as evidenced by the effective Fe3+ binding constants (K-eff') observed, which range from 1 x 10(17) M-1 to 4 x 10(18) M-1 at pH 6.5 and 20degreesC. The redox potentials for Fe(3+)nFbpX/Fee+nFbpX reduction are also found to depend on the identity of the synergistic anion required for Fe3+ sequestration. Facile exchange of exogenous anions (Fe(3+)nFbpX + X' --> Fe(3+)nFbpX' + X) is established and provides a pathway for environmental modulation of the iron chelation and redox characteristics of nFbp. The affinity of the iron loaded protein for exogenous anion binding at pH 6.5 was found to decrease in the order phosphate > arsenate similar to pyrophosphate > nitrilotriacetate > citrate similar to oxalate much greater than carbonate. Anion influence on the iron primary coordination sphere through iron binding and redox potential modulation may have in vivo application as a mechanism for periplasmic control of iron delivery to the cytosol.
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