Siderocalin (Scn), lipocalin-2, NGAL, 24p3 is a mammalian lipocalin-type protein that can prevent iron acquisition by pathogenic bacteria by binding siderophores, which are iron-binding chelators made by microorganisms. Iron serves as a key nutrient in host-pathogen interactions, and pathogens can acquire iron from the host organism via synthesis and release siderophores such as enterobactin. Siderocalin is a part of the mammalian defence mechanism and acts as an antibacterial agent. Crystallographic studies of Scn demonstrated that it includes a calyx, a ligand-binding domain that is lined with polar cationic groups. Central to the siderophore/siderocalin recognition mechanism are hybrid electrostatic/cation-pi interactions. To evade the host defences, pathogens evolved to produce structurally varied siderophores that would not be recognized by siderocalin, allowing the bacteria to acquire iron.
Iron requirements of host organisms Organisms require iron for a variety of chemical reactions. Although iron can be found throughout the biosphere, free ferric iron forms insoluble hydroxides at physiological pH, limiting its accessibility in aerobic conditions to living organisms. In order to preserve homeostasis, organisms have evolved specific protein networks, with proteins and receptors translated in accordance with intracellular iron levels. Export and import are supplemented by a cycling process between the ferrous Fe(II) available in the reducing environment of the cell, and ferric Fe(III) found primarily under aerobic conditions. The iron acquisition mechanisms of pathogenic bacteria demonstrate the role of iron as a key component at the interface between pathogens and hosts.
Lipocalin family of iron binding proteins The lipocalin family of binding proteins are produced by the immune system and sequester ferric siderophore complexes from the siderophore receptors of bacteria. The lipocalin family of binding proteins typically have a conserved eight-stranded β-barrel fold with a calyx binding site, which are lined with positively charged amino acid residues, allowing for binding interactions with siderophores.
Clinical significance
Mycobacterial infections The lipocalin siderocalin is found in neutrophil granules, uterine secretions, and at particularly high levels in serum during bacterial infection. Upon infection, pathogens use siderophores to capture iron from the host organism. This strategy is, however, complicated by the human protein siderocalin, which can sequester siderophores, and prevent their use by pathogenic bacteria as iron delivery agents. This effect has been demonstrated by studies with siderocalin-knock-out mice, which are more sensitive to infections under iron-limiting conditions.
Mycobacterial virulence Siderophores are iron chelators, allowing organisms to acquire iron from their environment. In the case of pathogens, iron can be acquired from the host organism. Siderophores and ferric iron can associate to form stable complexes. Siderophores bind iron using a variety of ligands, most commonly as α-hydroxycarboxylates (e.g. citrate), catecholates, and hydroxamates. As a defence mechanism, siderocalin can substitute ferric bis-catechol complexes (formed under physiological conditions) with a third catechol, in order to achieve a hexacoordinate ferric complex, resulting in higher affinity binding.
As a mediator of mammalian iron transport Mammalian siderophores, specifically catechols, can be found in the human gut and in siderophores, such as enterobactin, and serve as iron-binding moieties. Catechol resembling molecules can act as iron ligands in the cell and in systematic circulation, allowing siderocalin to bind to the iron-catechol complex. Catechols can be bound by siderocalin, in the form of free ligands, or in the iron complex. 24p3 is a vertebrate lipocalin-2 receptor which allows for import of the ferric siderophore complex into mammalian cells. During kidney embryogenesis, siderocalin mediated iron transport occurs, as iron concentration has to be highly controlled in order to restrict inflammation. Following secretion by neutrophils, siderocalin can bind to pathogenic siderophores, such as bacillibactin, and prevent siderophore trafficking. Siderocalin has been linked with various cellular processes apart from iron transport, including apoptosis, cellular differentiation, tumorigenesis, and metastasis.
Structure The avian orthologs of siderocalin (Q83 and Ex-FABP) and NGAL (neutrophil gelatinase-associated lipocalin-2) contain calyces with positively charged lysine and arginine side chains. These side chains interact via cation-pi and coulombic interactions with the negatively charged siderophores that contain aromatic catecholate groups. Crystallographic studies of siderocalin have shown that the ligand binding domain of Scn, known as the calyx, is shallow and broad, and is lined with polar cationic groups from the three positively charged residues of Arg81, Lys125, and Lys134. Scn can also bind non-ferric complexes and has been identified as a potential transporter for heavy actinide ions. Scn crystal structures containing heavy metals (thorium, plutonium, americium, curium, and californium) have been obtained. Scn has been found as a monomer, homo-dimer, or trimer in human plasma. The siderocalin fold is exceptionally stable. The calyx is structurally stable and rigid, and conformational change does not typically occur upon a change in pH, ionic strength, or ligand binding.
Binding pocket The structural stability of the calyx has been attributed to the three binding pockets within the calyx that sterically limit which ligands are compatible with siderocalin. The Scn calyx can accommodate three aromatic rings of the catecholate moieties, in the three available binding pockets. Solid-state and solution structural results demonstrated that bacteria-derived enterobactin is bound to the binding pocket of Scn, allowing for Scn to be involved in the acute immune response to bacterial infection. One method by which pathogens can circumvent immunity mechanisms is by modifying the siderophore chemical structure to prevent interaction with Scn. One example is the addition of glucose molecules to the enterobactin backbone of salmochelin (C-glucosylated enterobactin) in order to increase the hydrophilicity and bulkiness of a siderophore and inhibit binding to Scn.
Binding interactions
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