A symmetrical tetramer for S. aureus pyruvate carboxylase in complex with coenzyme A

Structure. 2009 Jun 10;17(6):823-32. doi: 10.1016/j.str.2009.04.008.

Abstract

Pyruvate carboxylase (PC) is a conserved metabolic enzyme with important cellular functions. We report crystallographic and cryo-electron microscopy (EM) studies of Staphylococcus aureus PC (SaPC) in complex with acetyl-CoA, an allosteric activator, and mutagenesis, biochemical, and structural studies of the biotin binding site of its carboxyltransferase (CT) domain. The disease-causing A610T mutation abolishes catalytic activity by blocking biotin binding to the CT active site, and Thr908 might play a catalytic role in the CT reaction. The crystal structure of SaPC in complex with CoA reveals a symmetrical tetramer, with one CoA molecule bound to each monomer, and cryo-EM studies confirm the symmetrical nature of the tetramer. These observations are in sharp contrast to the highly asymmetrical tetramer of Rhizobium etli PC in complex with ethyl-CoA. Our structural information suggests that acetyl-CoA promotes a conformation for the dimer of the biotin carboxylase domain of PC that might be catalytically more competent.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetyl Coenzyme A / metabolism*
  • Allosteric Regulation
  • Amino Acid Sequence
  • Binding Sites
  • Biotin / metabolism
  • Carbon-Nitrogen Ligases / chemistry
  • Carbon-Nitrogen Ligases / metabolism
  • Catalysis
  • Cryoelectron Microscopy
  • Crystallization
  • Crystallography, X-Ray
  • Dimerization
  • Enzyme Activation
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Quaternary
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Pyruvate Carboxylase / chemistry*
  • Pyruvate Carboxylase / genetics
  • Pyruvate Carboxylase / isolation & purification
  • Pyruvate Carboxylase / metabolism*
  • Pyruvate Carboxylase / ultrastructure
  • Staphylococcus aureus / enzymology*
  • X-Ray Diffraction

Substances

  • Biotin
  • Acetyl Coenzyme A
  • Carbon-Nitrogen Ligases
  • biotin carboxylase
  • Pyruvate Carboxylase