The role of electrostatics in siderophore recognition by the immunoprotein Siderocalin.
Iron is required for virulence of most bacterial pathogens, many of which rely on siderophores, small-molecule chelators, to scavenge iron in mammalian hosts. As an immune response, the human protein Siderocalin binds both apo and ferric siderophores in order to intercept delivery of iron to the bacterium, impeding virulence. The introduction of steric clashes into the siderophore structure is an important mechanism of evading sequestration. However, in the absence of steric incompatibilities, electrostatic interactions determine siderophore strength of binding by Siderocalin. By using a series of isosteric enterobactin analogues, the contribution of electrostatic interactions, including both charge-charge and cation-pi, to the recognition of 2,3-catecholate siderophores has been deconvoluted. The analogues used in the study incorporate a systematic combination of 2,3-catecholamide (CAM) and N-hydroxypyridinonate (1,2-HOPO) binding units on a tris(2-aminoethyl)amine (tren) backbone, [tren(CAM)(m)(1,2-HOPO)(n), where m = 0, 1, 2, or 3 and n = 3 - m]. The shape complementarity of the synthetic analogue series was determined through small-molecule crystallography, and the binding interactions were investigated through a fluorescence-based binding assay. These results were modeled and correlated through ab initio calculations of the electrostatic properties of the binding units. Although all the analogues are accommodated in the binding pocket of Siderocalin, the ferric complexes incorporating decreasing numbers of CAM units are bound with decreasing affinities (K(d) = >600, 43, 0.8, and 0.3 nM for m = 0-3). These results elucidate the role of electrostatics in the mechanism of siderophore recognition by Siderocalin.
Hoette
Trisha M
TM
Department of Chemistry, University of California, Berkeley, California 94720-1460, USA.
Abergel
Rebecca J
RJ
Xu
Jide
J
Strong
Roland K
RK
Raymond
Kenneth N
KN
D000818:Animals; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004095:Dihydropyridines / Q000737:chemistry*; D004758:Enterobactin / Q000737:chemistry; D006801:Humans; D007162:Immunoproteins / Q000737:chemistry; D007501:Iron / Q000737:chemistry; D007502:Iron Chelating Agents / Q000737:chemistry; D007700:Kinetics; D008024:Ligands; D008968:Molecular Conformation; D017262:Siderophores / Q000737:chemistry*; D012984:Software; D055672:Static Electricity; D013379:Substrate Specificity
D004095:Dihydropyridines; D007162:Immunoproteins; D007502:Iron Chelating Agents; D008024:Ligands; D017262:Siderophores; D004758:Enterobactin; D007501:Iron