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Formation of a protein intermediate and its trapping by the simultaneous crystallization process: Crystal structure of an iron-saturated intermediate in the FE3+ binding pathway of camel lactoferrin at 2.7 resolutionFormation of a protein intermediate and its trapping by the simultaneous crystallization process: Crystal structure of an iron-saturated intermediate in the FE3+ binding pathway of camel lactoferrin at 2.7 resolution
Structural highlights
Function[TRFL_CAMDR] Transferrins are iron binding transport proteins which can bind two Fe(3+) ions in association with the binding of an anion, usually bicarbonate (By similarity). The lactotransferrin transferrin-like domain 1 functions as a serine protease of the peptidase S60 family that cuts arginine rich regions. This function contributes to the antimicrobial activity (By similarity). Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedThis is the first protein intermediate obtained in the crystalline state by the simultaneous process of Fe(3+) binding and crystal nucleation and is also the first structure of an intermediate of lactoferrin in the Fe(3+) binding pathway. Lactoferrin is an iron-binding 80-kDa glycoprotein. It binds Fe(3+) very tightly in a closed interdomain cleft in both lobes. The iron-free structure of lactoferrin, on the other hand, adopts an open conformation with domains moving widely apart. These studies imply that initial Fe(3+) binding must be in the open form. The protein intermediate was crystallized by the microdialysis method. The protein solution, with a concentration of 100 mg/ml in 10 mm Tris-HCl, pH 8.0, was loaded in a capillary and dialyzed against the same buffer containing 26% (v/v) ethanol placed in a reservoir. FeCl(3) and CO(3)(2-) in excess molar ratios to that of protein in its solution were added to the reservoir buffer. The crystals appeared after some hours and grew to the optimum size within 36 h. The structure was determined by molecular replacement method and refined to final R- and R-free factors of 0.187 and 0.255, respectively. The present structure showed that the protein molecule adopts an open conformation similar to that of camel apolactoferrin. The electron density map clearly indicated the presence of two iron atoms, one in each lobe with 4-fold coordinations: two by the protein ligands of Tyr-92(433) OH and Tyr-192(526) OH and two other coordination sites occupied by oxygen atoms of bidentate CO(3)(2-) ions leading to a tetrahedral intermediate. The CO(3)(2-) anion is stabilized through hydrogen bonds with the synergistic anion-binding site Arg-121(463) and with Ser-122 Ogamma in the N-lobe and Thr-464 Ogamma in C-lobe. The third oxygen atom of CO(3)(2-) interacts with a water molecule in both lobes. Protein intermediate trapped by the simultaneous crystallization process. Crystal structure of an iron-saturated intermediate in the Fe3+ binding pathway of camel lactoferrin at 2.7 a resolution.,Khan JA, Kumar P, Srinivasan A, Singh TP J Biol Chem. 2001 Sep 28;276(39):36817-23. Epub 2001 Jul 25. PMID:11473113[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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