Identification and characterization of the heme-binding proteins SeShp and SeHtsA of Streptococcus equi subspecies equi.

BACKGROUND

Heme is a preferred iron source of bacterial pathogens. Streptococcus equi subspecies equi is a bacterial pathogen that causes strangles in horses. Whether S. equi has a heme acquisition transporter is unknown.

RESULTS

An S. equi genome database was blasted with the heme binding proteins Shp and HtsA of Streptococcus pyogenes, and found that S. equi has the homologue of Shp (designated SeShp) and HtsA (designated SeHtsA). Tag-free recombinant SeShp and SeHtsA and 6xHis-tagged SeHtsA (SeHtsAHis) were prepared and characterized. Purified holoSeShp and holoSeHtsA bind Fe(II)-protoporphyrin IX (heme) and Fe(III)-protoporphyrin IX (hemin) in a 1:1 stoichiometry, respectively, and are designated hemoSeShp and hemiSeHtsA. HemiSeShp and hemiSeHtsAHis can be reconstituted from apoSeShp and apoSeHtsAHis and hemin. HemoSeShp is stable in air and can be oxidized to hemiSeShp by ferricyanide. HemiSeHtsA can be reduced into hemoSeHtsA, which autoxidizes readily. HemoSeShp rapidly transfers its heme to apoSeHtsAHis. In addition, hemoSeShp can also transfer its heme to apoHtsA, and hemoShp is able to donate heme to apoSeHtsAHis.

CONCLUSION

The primary structures, optical properties, oxidative stability, and in vitro heme transfer reaction of SeShp and SeHtsA are very similar to those of S. pyogenes Shp and HtsA. The data suggest that the putative cell surface protein SeShp and lipoprotein SeHtsA are part of the machinery to acquire heme in S. equi. The results also imply that the structure, function, and functional mechanism of the heme acquisition machinery are conserved in S. equi and S. pyogenes.