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Iron-containing lipoprotein SiaA in SiaABC, the primary heme transporter of Streptococcus pyogenes

Abstract

The cell-surface lipoprotein SiaA, a component of the SiaABC transporter, acts as the primary receptor for heme in the infamous human pathogen Streptococcus pyogenes. However, little is known about the molecular mechanism of heme binding and release as well as the role of heme-binding ligands that contribute to the uptake of heme into the pathogenic bacteria. The present report aims to clarify the coordination properties of heme iron in SiaA. By substitution of either Met79 or His229 with alanine, the mutant M79A and H229A proteins display significantly decreased heme-binding affinity and substantially increased heme-release rates, as compared with wild-type SiaA protein. Both fluorescence and circular dichroism spectra indicated that heme binding results in alterations in the secondary structure of the protein. Heme release from SiaA is a stepwise process in which heme disassociates firstly from Met79 and then from His229 with distinct conformational changes. His229 may serve as an anchor for heme binding in SiaA and thus may play a major role in the stability of the coordination between heme and the protein.

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Abbreviations

ABC:

ATP-binding cassette

CD:

Circular dichroism

DTT:

Dithiothreitol

ICP-MS:

Inductively coupled plasma mass spectrometry

Ni-NTA:

Nickel nitrilotriacetic acid

rEK:

Recombinant enterokinase

Tris:

Tris(hydroxymethyl)aminomethane

WT:

Wild type

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Acknowledgments

This work was partially supported by the 2007 Chang-Jiang Scholars Program, “211” Projects, the National Natural Science Foundation of China (20871057, to Q.-Y. H.; 20801061, to R.G.), and Fundamental Research Funds for the Central Universities (to Q.-Y. H, and X. S.), and for Talents Start-Up (JNU 51208047, to X. S.).

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Correspondence to Qing-Yu He.

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Sun, X., Ge, R., Zhang, D. et al. Iron-containing lipoprotein SiaA in SiaABC, the primary heme transporter of Streptococcus pyogenes . J Biol Inorg Chem 15, 1265–1273 (2010). https://doi.org/10.1007/s00775-010-0684-4

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  • DOI: https://doi.org/10.1007/s00775-010-0684-4

Keywords

  • Streptococcus pyogenes
  • Heme-binding protein
  • Heme transport