Structural insight into the reaction mechanism and evolution of cytokinin biosynthesis

Proc Natl Acad Sci U S A. 2008 Feb 19;105(7):2734-9. doi: 10.1073/pnas.0707374105. Epub 2008 Feb 7.


The phytohormone cytokinin regulates plant growth and development. This hormone is also synthesized by some phytopathogenic bacteria, such as Agrobacterium tumefaciens, and is as a key factor in the formation of plant tumors. The rate-limiting step of cytokinin biosynthesis is catalyzed by adenosine phosphate-isopentenyltransferase (IPT). Agrobacterium IPT has a unique substrate specificity that enables it to increase trans-zeatin production by recruiting a metabolic intermediate of the host plant's biosynthetic pathway. Here, we show the crystal structures of Tzs, an IPT from A. tumefaciens, complexed with AMP and a prenyl-donor analogue, dimethylallyl S-thiodiphosphate. The structures reveal that the carbon-nitrogen-based prenylation proceeds by the SN2-reaction mechanism. Site-directed mutagenesis was used to determine the amino acid residues, Asp-173 and His-214, which are responsible for differences in prenyl-donor substrate specificity between plant and bacterial IPTs. IPT and the p loop-containing nucleoside triphosphate hydrolases likely evolved from a common ancestral protein. Despite structural similarities, IPT has evolved a distinct role in which the p loop transfers a prenyl moiety in cytokinin biosynthesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Monophosphate / chemistry
  • Adenosine Monophosphate / metabolism
  • Agrobacterium tumefaciens / chemistry
  • Agrobacterium tumefaciens / genetics
  • Agrobacterium tumefaciens / metabolism
  • Amino Acids / chemistry
  • Amino Acids / metabolism
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Binding Sites
  • Crystallography, X-Ray
  • Cytokinins / biosynthesis*
  • Cytokinins / chemistry
  • Evolution, Molecular*
  • Kinetics
  • Models, Molecular
  • Organothiophosphorus Compounds / chemistry
  • Organothiophosphorus Compounds / metabolism
  • Protein Binding
  • Protein Structure, Tertiary
  • Substrate Specificity
  • Zinc / chemistry
  • Zinc / metabolism


  • Amino Acids
  • Bacterial Proteins
  • Cytokinins
  • Organothiophosphorus Compounds
  • dimethyl thiophosphite
  • Adenosine Monophosphate
  • Zinc

Associated data

  • PDB/2ZE5
  • PDB/2ZE6
  • PDB/2ZE7
  • PDB/2ZE8