Open AccessIntracellular Metal Speciation in Streptococcus sanguinis Establishes SsaACB as Critical for Redox Maintenance
Author(s) -
Cody J. Murgas,
Shan P. Green,
Ashley K. Forney,
Rachel M. Korba,
Seon-Sook An,
Todd Kitten,
Heather R. Lucas
Publication year - 2020
Publication title -
acs infectious diseases
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.324
H-Index - 39
ISSN - 2373-8227
DOI - 10.1021/acsinfecdis.0c00132
Subject(s) - intracellular , streptococcus sanguinis , redox , virulence , chemistry , biophysics , siderophore , microbiology and biotechnology , biochemistry , biology , streptococcus mutans , bacteria , inorganic chemistry , genetics , gene
Streptococcus sanguinis is an oral commensal bacterium, but it can colonize pre-existing heart valve vegetations if introduced into the bloodstream, leading to infective endocarditis. Loss of Mn- or Fe-cofactored virulence determinants are thought to result in weakening of this bacterium. Indeed, intracellular Mn accumulation mediated by the lipoprotein SsaB, a component of the SsaACB transporter complex, has been shown to promote virulence for endocarditis and O 2 tolerance. To delineate intracellular metal-ion abundance and redox speciation within S. sanguinis , we developed a protocol exploiting two spectroscopic techniques, Inductively coupled plasma-optical emission spectrometry (ICP-OES) and electron paramagnetic resonance (EPR) spectroscopy, to respectively quantify total intracellular metal concentrations and directly measure redox speciation of Fe and Mn within intact whole-cell samples. Addition of the cell-permeable siderophore deferoxamine shifts the oxidation states of accessible Fe and Mn from reduced-to-oxidized, as verified by magnetic moment calculations, aiding in the characterization of intracellular metal pools and metal sequestration levels for Mn 2+ and Fe. We have applied this methodology to S. sanguinis and an SsaACB knockout strain (Δ ssaACB ), indicating that SsaACB mediates both Mn and Fe uptake, directly influencing the metal-ion pools available for biological inorganic pathways. Mn supplementation of Δ ssaACB returns total intracellular Mn to wild-type levels, but it does not restore wild-type redox speciation or distribution of metal cofactor availability for either Mn or Fe. Our results highlight the biochemical basis for S. sanguinis oxidative resistance, revealing a dynamic role for SsaACB in controlling redox homeostasis by managing the intracellular Fe/Mn composition and distribution.