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Crystal structure of the periplasmic component of a tripartite macrolide-specific efflux pump

Soohwan Yum1, Yongbin Xu, Shunfu Piao

  • 1College of Pharmacy and Research Institute for Drug Development, Pusan National University, Busan 609-735, Korea.

Journal of Molecular Biology
|March 4, 2009
PubMed

Insights

Researchers discovered that the MacA protein in Gram-negative bacteria forms a hexameric structure. This finding clarifies the oligomeric state of membrane fusion proteins (MFPs) in essential secretion and drug efflux systems.

Area of Science:

  • Microbiology
  • Structural Biology
  • Biochemistry

Background:

  • Type I protein secretion systems and tripartite drug efflux pumps in Gram-negative bacteria utilize a periplasmic membrane fusion protein (MFP).
  • MFPs are crucial for bridging outer membrane factors and inner membrane transporters, but their oligomeric state has been unclear.
  • AcrA is a well-characterized MFP in the AcrB-AcrA-TolC multidrug efflux pump of Escherichia coli.

Purpose of the Study:

  • To determine the oligomeric state and structure of the MacA protein, a periplasmic MFP.
  • To compare the structural features of MacA with other known MFPs like AcrA.
  • To elucidate the role of MacA's oligomeric assembly in the function of the MacAB-TolC pump.

Main Methods:

  • X-ray crystallography was used to determine the crystal structure of E. coli MacA and obtain an experimentally phased map of Actinobacillus actinomycetemcomitans MacA.
  • Transmission electron microscopy (TEM) was employed to visualize the MacA assembly.
  • In vitro and in vivo functional studies were conducted to confirm the biological relevance of the hexameric structure.

Main Results:

  • The crystal structures revealed a domain orientation in MacA distinct from AcrA.
  • Both crystal structures consistently showed MacA forming a hexameric assembly with a funnel-like shape, featuring a central channel and a conical mouth.
  • Electron microscopy and functional assays confirmed the hexameric nature of MacA and its importance in the MacAB-TolC pump.

Conclusions:

  • MacA adopts a hexameric conformation, clarifying the oligomeric state of this MFP.
  • The funnel-like hexameric structure of MacA provides structural insights into its function within drug efflux pumps and type I secretion systems.
  • This study advances the understanding of molecular mechanisms underlying bacterial transport systems.

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