Structure of the biotinyl domain of acetyl-coenzyme A carboxylase determined by MAD phasing

Structure. 1995 Dec 15;3(12):1407-19. doi: 10.1016/s0969-2126(01)00277-5.


Background: Acetyl-coenzyme A carboxylase catalyzes the first committed step of fatty acid biosynthesis. Universally, this reaction involves three functional components all related to a carboxybiotinyl intermediate. A biotinyl domain shuttles its covalently attached biotin prosthetic group between the active sites of a biotin carboxylase and a carboxyl transferase. In Escherichia coli, the three components reside in separate subunits: a biotinyl domain is the functional portion of one of these, biotin carboxy carrier protein (BCCP).

Results: We have expressed natural and selenomethionyl (Se-met) BCCP from E. coli as biotinylated recombinant proteins, proteolyzed them with subtilisin Carlsberg to produce the biotinyl domains BCCP and Se-met BCCPsc, determined the crystal structure of Se-met BCCPsc using a modified version of the multiwavelength anomalous diffraction (MAD) phasing protocol, and refined the structure for the natural BCCPsc at 1.8 A resolution. The structure may be described as a capped beta sandwich with quasi-dyad symmetry. Each half contains a characteristic hammerhead motif. The biotinylated lysin is located at a hairpin beta turn which connects the two symmetric halves of the molecule, and its biotinyl group interacts with a non-symmetric protrusion from the core.

Conclusions: This first crystal structure of a biotinyl domain helps to unravel the central role of such domains in reactions catalyzed by biotin-dependent carboxylases. The hammerhead structure observed twice in BCCPsc may be regarded as the basic structural motif of biotinyl and lipoyl domains of a superfamily of enzymes. The new MAD phasing techniques developed in the course of determining this structure enhance the power of the MAD method.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acetyl-CoA Carboxylase / chemistry*
  • Acetyl-CoA Carboxylase / metabolism
  • Amino Acid Oxidoreductases / chemistry
  • Amino Acid Sequence
  • Bacterial Proteins / chemistry*
  • Binding Sites
  • Biotin / chemistry*
  • Biotin / metabolism
  • Crystallography, X-Ray / methods*
  • Escherichia coli / enzymology*
  • Evolution, Molecular
  • Geobacillus stearothermophilus / enzymology
  • Glycine Dehydrogenase (Decarboxylating)
  • Humans
  • Hydrogen Bonding
  • Lysine / analogs & derivatives
  • Lysine / chemistry
  • Lysine / metabolism
  • Models, Molecular*
  • Molecular Sequence Data
  • Multiple Carboxylase Deficiency / enzymology
  • Peptide Fragments / chemistry*
  • Plant Proteins / chemistry
  • Protein Binding
  • Protein Structure, Secondary
  • Protein Structure, Tertiary*
  • Pyruvate Dehydrogenase Complex / chemistry
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Proteins*
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Species Specificity
  • Structure-Activity Relationship


  • BCCPsc protein, recombinant
  • Bacterial Proteins
  • Peptide Fragments
  • Plant Proteins
  • Pyruvate Dehydrogenase Complex
  • Recombinant Fusion Proteins
  • Recombinant Proteins
  • Se-Met-BCCPsc protein, recombinant
  • Biotin
  • Amino Acid Oxidoreductases
  • Glycine Dehydrogenase (Decarboxylating)
  • Acetyl-CoA Carboxylase
  • biocytin
  • Lysine