Kinetics of iron release from ferric binding protein (FbpA): mechanistic implications in bacterial periplasm-to-cytosol Fe3+ transport.
The ferric binding protein (FbpA) transports iron across the periplasmic space of certain Gram-negative bacteria and is an important component involved in iron acquisition by pathogenic Neisseria spp. (Neisseria gonorrheae and Neisseria meningitidis). Previous work has demonstrated that the synergistic anion, required for tight Fe(3+) sequestration by FbpA, also plays a key role in inserting Fe(3+) into the FbpA binding site. Here, we investigate the iron release process from various forms of holo-FbpA, Fe(3+)FbpA-X, during the course of a chelator competition reaction using EDTA and Tiron. Fe(3+)FbpA-X represents the protein assembly complex with different synergistic anions, X = PO(4)(3)(-) and NTA. Stepwise mechanisms of Fe(3+) release are proposed on the basis of kinetic profiles of these chelator competition reactions. Fe(3+)FbpA-PO(4) and Fe(3+)FbpA-NTA react differently with EDTA and Tiron during the Fe(3+)-exchange process. EDTA replaces PO(4)(3)(-) and NTA from the first coordination shell of Fe(3+) and acts as a synergistic anion to give a spectroscopically distinguishable intermediate, Fe(3+)FbpA-EDTA, prior to pulling Fe(3+) out of the protein. Tiron, on the other hand, does not act as a synergistic anion but is a more efficient competing chelator as it removes Fe(3+) from FbpA at rate much faster than EDTA. These results reaffirm the contribution of the synergistic anion to the FbpA iron transport process as the anion, in addition to playing a facilitative role in iron binding, appears to have a "gatekeeper" role, thereby modulating the Fe(3+) release process.
Dhungana
Suraj
S
Department of Chemistry, Duke University, Durham, North Carolina 27708-0346, USA.
Anderson
Damon S
DS
Mietzner
Timothy A
TA
Crumbliss
Alvin L
AL
D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001693:Biological Transport, Active; D027682:Cation Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D003600:Cytosol / Q000737:chemistry* / Q000378:metabolism; D004357:Drug Synergism; D004492:Edetic Acid / Q000737:chemistry; D005290:Ferric Compounds / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007501:Iron / Q000737:chemistry* / Q000378:metabolism; D007502:Iron Chelating Agents / Q000737:chemistry; D033862:Iron-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008956:Models, Chemical; D009343:Neisseria / Q000737:chemistry / Q000378:metabolism; D009571:Nitrilotriacetic Acid / Q000737:chemistry; D019897:Periplasm / Q000737:chemistry* / Q000378:metabolism; D013053:Spectrophotometry; D013816:Thermodynamics; D014168:Transferrin / Q000737:chemistry / Q000378:metabolism
D001426:Bacterial Proteins; D027682:Cation Transport Proteins; D005290:Ferric Compounds; D007502:Iron Chelating Agents; D033862:Iron-Binding Proteins; D014168:Transferrin; C471557:iron-transport protein, Neisseria gonorrhoeae; D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; D004492:Edetic Acid; D007501:Iron; D009571:Nitrilotriacetic Acid