Mutagenesis, biochemical, and biophysical characterization of Mycoplasma arthritidis-derived mitogen

Hongmin Li, Yiwei Zhao, Yi Guo, Sandra J. VanVranken, Zhong Li, Leslie Eisele, Walid Mourad

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

Mycoplasma arthritidis-derived mitogen (MAM) is a superantigen (SAg) that can activate large fractions of T cells bearing particular TCR Vβ elements. Here we report the mutagenesis, biochemical and biophysical studies on the dimerization of MAM in solution. Our studies showed that although MAM mainly exists as a monomer in solution, a small percentage of MAM molecules form homodimer at high protein concentration, regardless of the presence of Zn2+. A distinct peak corresponding to a MAM homodimer was detected in the presence of EDTA, using both chemical cross-linking and analytical ultracentrifugation methods. Further mutagenesis studies revealed that single mutation of residues at the interface of the crystallographic dimer of MAM does not significantly affect the dimerization of MAM in solution. Circular dichroism (CD) analysis indicated that addition of Zn2+ does not induce conformational changes of MAM from its apo-state. Thermal denaturation experiments indicated that addition of Zn2+ to MAM solution resulted in a decrease of melting point (Tm), whereas addition of EDTA did not affect the Tm of MAM. These results imply that there is no defined Zn2+-binding site on MAM.

Original languageEnglish (US)
Pages (from-to)763-773
Number of pages11
JournalMolecular Immunology
Volume44
Issue number5
DOIs
StatePublished - Feb 2007
Externally publishedYes

Keywords

  • Analytical ultracentrifugation
  • CD
  • Cross-linking
  • Dimerization
  • MAM
  • Sedimentation
  • Superantigen

ASJC Scopus subject areas

  • Immunology
  • Molecular Biology

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