Extracellular Zinc Competitively Inhibits Manganese Uptake and Compromises Oxidative Stress Management in Streptococcus pneumoniae

Abstract

Streptococcus pneumoniae requires manganese for colonization of the human host, but the underlying molecular basis for this requirement has not been elucidated. Recently, it was shown that zinc could compromise manganese uptake and that zinc levels increased during infection by S. pneumoniae in all the niches that it colonized. Here we show, by quantitative means, that extracellular zinc acts in a dose dependent manner to competitively inhibit manganese uptake by S. pneumoniae, with an EC50 of 30.2 µM for zinc in cation-defined media. By exploiting the ability to directly manipulate S. pneumoniae accumulation of manganese, we analyzed the connection between manganese and superoxide dismutase (SodA), a primary source of protection for S. pneumoniae against oxidative stress. We show that manganese starvation led to a decrease in sodA transcription indicating that expression of sodA was regulated through an unknown manganese responsive pathway. Intriguingly, examination of recombinant SodA revealed that the enzyme was potentially a cambialistic superoxide dismutase with an iron/manganese cofactor. SodA was also shown to provide the majority of protection against oxidative stress as a S. pneumoniae ΔsodA mutant strain was found to be hypersensitive to oxidative stress, despite having wild-type manganese levels, indicating that the metal ion alone was not sufficiently protective. Collectively, these results provide a quantitative assessment of the competitive effect of zinc upon manganese uptake and provide a molecular basis for how extracellular zinc exerts a ‘toxic’ effect on bacterial pathogens, such as S. pneumoniae.

Overlapping Functionality of the Pht Proteins in Zinc Homeostasis of Streptococcus pneumoniae

“…The results of this work and prior studies support the involvement of the Pht proteins in facilitating Zn 2ϩ acquisition from the environment under conditions where its availability is tightly restricted (9). However, it is also plausible that they could help defend the pneumococcus when an excess of the metal ion is present in the extracellular environment, which exerts a toxic effect on the bacteria by interfering with the physiological function of the ABC permease, PsaBCA, in importing the essential transition metal ion manganese (20,21). It has been speculated that the Pht proteins may aid in protecting the PsaBCA permease from extracellular Zn 2ϩ by binding excess ions and acting as a "sink" (12,22).…”

The Role of Copper and Zinc Toxicity in Innate Immune Defense against Bacterial Pathogens

“…However, like Cu, Zn lies at the top of the Irving-Williams series, and there is evidence that Zn exerts an antimicrobial effect by antagonizing the uptake of other key trace metal nutrients. In Streptococcus pneumoniae, excess Zn was shown to prevent the uptake of Mn(II) by binding irreversibly to the extracellular Mn(II) solute binding protein PsaA (45,46). Consequently, S. pneumoniae becomes hypersensitive to oxidative stress (45,46).…”

“…In Streptococcus pneumoniae, excess Zn was shown to prevent the uptake of Mn(II) by binding irreversibly to the extracellular Mn(II) solute binding protein PsaA (45,46). Consequently, S. pneumoniae becomes hypersensitive to oxidative stress (45,46). Zn may also bind adventitiously to other sites that do not normally contain a metal ion and thereby inhibit key enzymes in cells (35,(47)(48)(49).…”

Functional Determinants of Metal Ion Transport and Selectivity in Paralogous Cation Diffusion Facilitator Transporters CzcD and MntE in Streptococcus pneumoniae

“…Excess iron (Fe) is harmful due to its participation in Fenton chemistry, which produces a highly reactive hydroxyl radical that can damage biomolecules (4-7). Other transition metals, including Mn, Zn, cobalt (Co), nickel (Ni), and copper (Cu), become toxic at high concentrations partly because they compete for each other's cognate metal-binding sites in enzymes (8)(9)(10)(11)(12)(13)(14)(15)(16)(17). Regardless of the mechanism of metal toxicity, bacteria have evolved ways to minimize the deleterious impact of metal ion excess.…”