Biochemical and Spectroscopic Observation of Mn(II) Sequestration from Bacterial Mn(II) Transport Machinery by Calprotectin

Abstract

Human calprotectin (CP, S100A8/S100A9 oligomer) is a metal-sequestering host-defense protein that prevents bacterial acquisition of Mn(II). In this work, we investigate Mn(II) competition between CP and two solute-binding proteins that Staphylococcus aureus and Streptococcus pneumoniae, Gram-positive bacterial pathogens of significant clinical concern, use to obtain Mn(II) when infecting a host. Biochemical and electron paramagnetic resonance (EPR) spectroscopic analyses demonstrate that CP outcompetes staphylococcal MntC and streptococcal PsaA for Mn(II). This behavior requires the presence of excess Ca(II) ions, which enhance the Mn(II) affinity of CP. This report presents new spectroscopic evaluation of two Mn(II) proteins important for bacterial pathogenesis, direct observation of Mn(II) sequestration from bacterial Mn(II) acquisition proteins by CP, and molecular insight into the extracellular battle for metal nutrients that occurs during infection.

Synergy between Nutritional Immunity and Independent Host Defenses Contributes to the Importance of the MntABC Manganese Transporter during Staphylococcus aureus Infection

“…Notably, loss of MntABC did not reduce the ability of S. aureus to obtain any of the other assayed metals. Collectively, these results indicate that, despite CP having a greater affinity for Mn in vitro (45), MntABC is necessary, as it enables S. aureus to obtain sufficient (but not optimal) quantities of Mn during infection. Further, our work also suggested that pathogens that express MntABC homologs compete more effectively with the host for Mn than those that express only homologs of MntH.…”

“…S. aureus possesses two Mn transporters, MntABC and MntH (12,27,28), and loss of MntABC has been shown to reduce staphylococcal virulence during infection (27,42,43). However, knowledge of the molecular basis for this defect has remained elusive, and recent work has suggested that MntABC may not be the primary transporter responsible for competing with the host for Mn (45). The current investigation revealed that MntABC is the primary transporter responsible for resisting host-imposed Mn limitation and that additive insults of nutritional immunity and other host defenses drive the importance of this transporter during infection.…”

“…In vitro studies have suggested near-parity between MntABC and MntH with respect to their contributions to growth in Mn-limited environments. However, binding studies have shown that CP can outcompete MntC for Mn (45), suggesting that MntABC might not be the primary staphylococcal Mn transporter during infection. Despite this, multiple animal models have shown that MntABC has a critical role in staphylococcal pathogenesis (27,42,43).…”

“…Although in vitro growth defects associated with loss of MntABC have been reported, those previous studies investigated conditions that are presumed to represent metal-replete environments (42,44). As CP has a greater affinity for Mn than MntC (45), this raises the possibility that MntABC may not be the primary transporter responsible for resisting host-imposed Mn starvation. Therefore, the relative contributions of MntABC and MntH to resisting nutritional immunity remain unclear.…”

Biophysical Examination of the Calcium-Modulated Nickel-Binding Properties of Human Calprotectin Reveals Conformational Change in the EF-Hand Domains and His3Asp Site

“…1–5 CP exhibits broad-spectrum antimicrobial activity attributed to its ability to sequester essential metal nutrients from invading pathogens. 1,4,6,7 Human CP coordinates divalent first-row transition metals, including Mn(II), 8–13 Fe(II), 14–16 Ni(II), 17 Cu(II), 18 and Zn(II), 8,19,20 with sufficiently high affinity to prevent microbial acquisition of these nutrients.…”