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Streptococcus sanguinis, present in the biofilm colonizing human tooth surfaces, may be beneficial in the oral cavity, though it also serves as a causative agent of an extra-oral disease, infective endocarditis. Previous mutagenesis of lipoprotein genes from S. sanguinis strain SK36 identified the SsaB gene as necessary for virulence in a rabbit model of endocarditis. Based on homology, we suspected that SsaB was a metal transporter. A procedure utilizing inductively coupled plasma optical emission spectrometry was optimized and implemented for determination of the concentrations (μg metal per mg of cellular protein) of 17 metals in SK36 and ssaB mutant cell lysates. Data revealed that the ssaB mutant reproducibly accumulated lower concentrations of both manganese and iron, but showed no significant difference in accumulated concentrations of magnesium, or any of 14 additional metals for which there were standards, or in signal intensities for 60 other elements analyzed for which there were no standards. This data suggested that SsaB binds both manganese and iron to facilitate transport. When grown in brain-heart infusion (BHI) broth, both strains accumulated more iron than manganese. To assess whether this was due entirely to the higher concentration of iron relative to manganese in BHI broth, SK36 and the ssaB mutant were grown in all-purpose tween (APT) broth, rich in both metals. Both strains accumulated more manganese than iron in APT broth. Higher accumulation of manganese in APT broth compared to BHI broth for both strains suggested that manganese is accumulated through both SsaB-dependent and SsaB-independent mechanisms. Relative cellular abundance of iron and manganese in S. sanguinis varies dramatically depending on relative abundance in the growth medium, highlighting the importance of using physiologically relevant media in future studies. This data also implies that S. sanguinis is flexible in its metal requirements and is rather efficient in sequestering iron, which would otherwise react with cellular hydrogen peroxide to produce DNA-damaging hydroxyl radicals.

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Todd Kitten


Virginia Commonwealth University. Undergraduate Research Opportunities Program

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Characterization of Metal Transport by the Streptococcus sanguinis Endocarditis Virulence Factor SsaB