Structure-function analysis of a bacterial deoxyadenosine kinase reveals the basis for substrate specificity

J Mol Biol. 2007 Mar 9;366(5):1615-23. doi: 10.1016/j.jmb.2006.12.010. Epub 2006 Dec 8.

Abstract

Deoxyribonucleoside kinases (dNKs) catalyze the transfer of a phosphoryl group from ATP to a deoxyribonucleoside (dN), a key step in DNA precursor synthesis. Recently structural information concerning dNKs has been obtained, but no structure of a bacterial dCK/dGK enzyme is known. Here we report the structure of such an enzyme, represented by deoxyadenosine kinase from Mycoplasma mycoides subsp. mycoides small colony type (Mm-dAK). Superposition of Mm-dAK with its human counterpart's deoxyguanosine kinase (dGK) and deoxycytidine kinase (dCK) reveals that the overall structures are very similar with a few amino acid alterations in the proximity of the active site. To investigate the substrate specificity, Mm-dAK has been crystallized in complex with dATP and dCTP, as well as the products dCMP and dCDP. Both dATP and dCTP bind to the enzyme in a feedback-inhibitory manner with the dN part in the deoxyribonucleoside binding site and the triphosphates in the P-loop. Substrate specificity studies with clinically important nucleoside analogs as well as several phosphate donors were performed. Thus, in this study we combine structural and kinetic data to gain a better understanding of the substrate specificity of the dCK/dGK family of enzymes. The structure of Mm-dAK provides a starting point for making new anti bacterial agents against pathogenic bacteria.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Amino Acids
  • Bacterial Proteins / analysis
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / isolation & purification
  • Bacterial Proteins / metabolism
  • Binding Sites
  • Conserved Sequence
  • Crystallography, X-Ray
  • Deoxyadenine Nucleotides / metabolism
  • Deoxycytosine Nucleotides / metabolism
  • Dimerization
  • Drug Design
  • Humans
  • Hydrogen Bonding
  • Ligands
  • Models, Molecular
  • Molecular Sequence Data
  • Molecular Weight
  • Mycoplasma mycoides / enzymology
  • Phosphotransferases (Alcohol Group Acceptor) / analysis
  • Phosphotransferases (Alcohol Group Acceptor) / chemistry*
  • Phosphotransferases (Alcohol Group Acceptor) / isolation & purification
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism*
  • Protein Binding
  • Protein Structure, Secondary
  • Sequence Homology, Amino Acid
  • Structure-Activity Relationship
  • Substrate Specificity

Substances

  • Amino Acids
  • Bacterial Proteins
  • Deoxyadenine Nucleotides
  • Deoxycytosine Nucleotides
  • Ligands
  • 2'-deoxycytidine 5'-triphosphate
  • Phosphotransferases (Alcohol Group Acceptor)
  • deoxyadenosine kinase
  • 2'-deoxyadenosine triphosphate

Associated data

  • PDB/2JAQ
  • PDB/2JAS
  • PDB/2JAT