Identification of possible kinetically significant anion‐binding sites in human serum transferrin using molecular modeling strategies

Abstract

Certain anions have been shown experimentally to influence the rate of iron release from human serum transferrin (HST), implying the existence of one or more allosteric kinetically significant anion-binding (KISAB) sites on or near the surface of the protein. A rank-ordered selection of potential HST KISAB sites has been obtained using a novel three-stage molecular modeling strategy. The crystal structure of HST (1A8E.pdb) was first subjected to a heuristic analysis, in which positively charged and hydrogen-bonding residues on or near the surface of the protein were identified. In this stage, a preliminary electrostatic potential map was also calculated, yielding six preliminary sites. Next, energy-grid calculations were conducted in order to identify anion-protein interaction energy minima, which resulted in the inclusion of three additional sites. Finally, three anions already shown experimentally to demonstrate varied effects on HST iron-release kinetics were placed at each potential site; molecular dynamics and molecular mechanics calculations were performed in order to elucidate the hydrogen-bonding environment around each anion of the protein as well as to calculate anion-protein-binding energies.

Identification of a Kinetically Significant Anion Binding (KISAB) Site in the N-Lobe of Human Serum Transferrin

“…We suggest that the absence of Lys144 might increase access of chloride ions to Arg143, resulting in an enhancement of iron release relative to the control. Although both Arg143 and Lys144 were suggested as a potential KISAB sites in a modeling study (42), our data clearly show that, unlike Arg143, Lys144 does not meet the criteria for this designation.…”

Molecular Dynamics Study of Naturally Existing Cavity Couplings in Proteins

“…To check whether our CSCs overlapped with allosteric couplings, and to which extent, we explored the literature and the Allosteric Database[ 72 ] (ASD, version 2.0). The information retrieved shows that experimental data were available for nine proteins: aldolase[ 73 , 74 ], annexin V[ 75 – 78 ], transferrin[ 79 – 82 ], retinol-binding protein[ 83 – 86 ] (RBP), purine nucleoside phosphorylase[ 87 , 88 ] (PNP), heat shock protein HSP90[ 89 ], cyclophilin A[ 90 – 92 ], interleukin-1alpha[ 93 , 94 ], and Awd nucleotide diphosphate kinase[ 95 ]. When comparing functional site (FS)/allosteric site (AS) annotations with our CSC data, we found several cases where the regions linked by both couplings overlapped: one cavity in the CSC would involve a number of FS residues and the other cavity would involve a number of AS residues (see below).…”

Release of iron from transferrin by phosphonocarboxylate and diphosphonate chelating agents

“…This high anion-binding affinity is consistent with the hypothesis that these anionic ligands can increase the value of k max by binding to an allosteric anion-binding site in the protein. Computational studies have identified a number of potential anion-binding sites in the Nlobe of ferric-transferrin [46]. However, it must be stressed that the binding constants in Table 4 are for apotransferrin, not ferric transferrin.…”