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FAD binding by ApbE protein from Salmonella enterica: a new class of FAD-binding proteins

Jeffery M Boyd1, James A Endrizzi, Trinity L Hamilton

  • 1Department of Bacteriology, University of Wisconsin, Madison, WI 53706, USA.

Journal of Bacteriology
|December 15, 2010
PubMed

Insights

The periplasmic protein ApbE binds flavin adenine dinucleotide (FAD). Mutagenesis identified a key tyrosine residue essential for ApbE

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Enzymology

Background:

  • The periplasmic protein ApbE is involved in thiamine biosynthesis and iron-sulfur cluster metabolism.
  • Mutations in apbE lead to reduced activity of iron-sulfur enzymes, but ApbE's specific function remains unclear.
  • Related proteins, NosX and RnfF, are implicated in redox processes in nitrogen metabolism.

Purpose of the Study:

  • To elucidate the molecular function of the periplasmic protein ApbE.
  • To investigate the interaction of ApbE with flavin adenine dinucleotide (FAD).
  • To identify key residues involved in FAD binding and protein function.

Main Methods:

  • Biochemical assays to determine FAD binding stoichiometry.
  • X-ray crystallography to determine the structure of ApbE with bound FAD.
  • Random and targeted mutagenesis to generate nonfunctional ApbE variants.
  • Reconstitution and FAD binding analysis of mutated ApbE proteins.

Main Results:

  • ApbE binds one molecule of FAD per monomer.
  • The crystal structure revealed a novel FAD-binding motif within ApbE.
  • A specific tyrosine residue mutation abolished FAD binding and protein reconstitution.

Conclusions:

  • ApbE is an FAD-binding protein with a unique FAD-binding motif.
  • The identified tyrosine residue is critical for ApbE's FAD interaction and likely its function.
  • These findings provide insights into the role of ApbE in cellular processes requiring iron-sulfur clusters.

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