A novel NAD-binding protein revealed by the crystal structure of 2,3-diketo-L-gulonate reductase (YiaK)

J Biol Chem. 2004 Mar 26;279(13):13148-55. doi: 10.1074/jbc.M313580200. Epub 2004 Jan 12.


Escherichia coli YiaK catalyzes the reduction of 2,3-diketo-L-gulonate in the presence of NADH. It belongs to a large family of oxidoreductases that is conserved in archaea, bacteria, and eukaryotes but shows no sequence homology to other proteins. We report here the crystal structures at up to 2.0-A resolution of YiaK alone and in complex with NAD-tartrate. YiaK has a new polypeptide backbone fold and a novel mode of recognizing the NAD cofactor. In addition, NAD is bound in an unusual conformation, at the interface of a dimer of the enzyme. The crystallographic analysis unexpectedly revealed the binding of tartrate in the active site. Enzyme kinetics studies confirm that tartrate and the related D-malate are inhibitors of YiaK. In contrast to most other enzymes where substrate binding produces a more closed conformation, the binding of NAD-tartrate to YiaK produces a more open active site. The free enzyme conformation is incompatible with NAD binding. His(44) is likely the catalytic residue of the enzyme.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Catalytic Domain
  • Crystallography, X-Ray
  • Dimerization
  • Electrons
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / metabolism
  • Kinetics
  • Models, Chemical
  • Models, Molecular
  • Molecular Sequence Data
  • NAD / chemistry*
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid
  • Sugar Acids / chemistry*
  • Sugar Alcohol Dehydrogenases / chemistry*
  • Sugar Alcohol Dehydrogenases / metabolism
  • Tartrates / chemistry


  • Escherichia coli Proteins
  • Sugar Acids
  • Tartrates
  • NAD
  • gulonic acid
  • Sugar Alcohol Dehydrogenases
  • 2,3-diketo-L-gulonate reductase
  • tartaric acid

Associated data

  • PDB/1NXU
  • PDB/1S20