Human calprotectin is an iron-sequestering host-defense protein

Abstract

Human calprotectin (CP) is a metal-chelating antimicrobial protein of the innate immune response. The current working model states that CP sequesters manganese and zinc from pathogens. We report the discovery that CP chelates iron and deprives bacteria of this essential nutrient. Elemental analysis of CP-treated growth medium establishes that CP reduces the concentrations of manganese, iron, and zinc. Microbial growth studies reveal that iron depletion by CP contributes to the growth inhibition of bacterial pathogens. Biochemical investigations demonstrate that CP coordinates Fe(II) at an unusual hexahistidine motif, and the Mössbauer spectrum of 57Fe(II)-bound CP is consistent with coordination of high-spin Fe(II) at this site (δ = 1.20 mm/s, ΔEQ = 1.78 mm/s). In the presence of Ca(II), CP turns on its iron-sequestering function and exhibits sub-picomolar affinity for Fe(II). Our findings expand the biological coordination chemistry of iron and support a previously unappreciated role for CP in mammalian iron homeostasis.

Salivary metals, age, and gender correlate with cultivable oral Candida carriage levels

“…As both calprotectin and Histatin 5 bind multiple different metals [14,17] related to their antimicrobial activity, this may reflect early host innate immune responses to higher Candida carriage. Of particular interest was our finding of the robust interaction between carriage with manganese and copper.…”

“…Saliva contains many proteins involved in nutritional immunity [5,10,12]. Calprotectin is the classic example of a metal-sequestering protein [10] that binds manganese, zinc, and iron with nanomolar to picomolar affinity, and inhibits microbial growth by disrupting superoxide dismutase function [13,14]. Salivary amylase [15], ferritin [16], Histatin 5 [17], gustin [18], and lactoferrin (functioning synergistically with calprotectin) [19] are all metal binding proteins expected to have a role in nutritional immunity.…”

Role of Calprotectin in Withholding Zinc and Copper from Candida albicans

“…To directly test whether CP is depleting available metals from the growth medium, we adapted the method of Nakashige et al (21), whereby CP-bound metals are removed by filtration. As seen in Fig.…”

“…CP has been well studied for its Zn-withholding properties from various pathogenic organisms, including Gram-positive and Gramnegative bacteria as well as the fungus Aspergillus fumigatus, where a mutant deficient in Zn sensing shows greater susceptibility toward CP in vivo (6,(13)(14)(15)(16)(17)(18). In addition to Zn, CP can withhold Mn from bacterial pathogens in vitro and in vivo (13,14,16,19,20) and can deplete Fe and Ni from specific microbial cultures in vitro (21,22). CP comprises 40 to 60% of the total cytoplasmic protein of neutrophils, accumulates at sites of infection at concentrations of up to 1 mg ml Ϫ1 (23,24), and is an important component of neutrophil extracellular traps (NETs) released in the extracellular space during a unique form of neutrophil cell death (25,26).…”

“…CP is a heterodimeric protein consisting of S100A8 and S100A9 subunits and acts by binding to extracellular metals using two distinct metal coordinating sites (27). Site 1 (S1) consists of six histidine residues, two from S100A8 and four from S100A9 (28,29), and has long been known to bind Zn(II) and Mn(II) (28)(29)(30)(31)(32)(33) and more recently known to bind Fe(II) and Ni(II) (21,22). Site 2 (S2) contains a canonical S100 transition metal binding site consisting of two histidines from S100A8 and a histidine and aspartate from S100A9.…”