title: Structural insight into the reaction mechanism and evolution of cytokinin biosynthesis.
abstract: 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.
authors.affiliation: RIKEN Plant Science Center, 1-7-22 Suehiro, Tsurumi, Yokohama 230-0045, Japan.
authors.forename: Hajime
authors.initials: H
authors.lastname: Sugawara
authors.forename: Nanae
authors.initials: N
authors.lastname: Ueda
authors.forename: Mikiko
authors.initials: M
authors.lastname: Kojima
authors.forename: Nobue
authors.initials: N
authors.lastname: Makita
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Yamaya
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Sakakibara
mesh_terms: D000249:Adenosine Monophosphate / Q000737:chemistry / Q000378:metabolism; D016960:Agrobacterium tumefaciens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D018360:Crystallography, X-Ray; D003583:Cytokinins / Q000096:biosynthesis* / Q000737:chemistry; D019143:Evolution, Molecular; D007700:Kinetics; D008958:Models, Molecular; D009946:Organothiophosphorus Compounds / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D015032:Zinc / Q000737:chemistry / Q000378:metabolism
chemical_list: D000596:Amino Acids; D001426:Bacterial Proteins; D003583:Cytokinins; D009946:Organothiophosphorus Compounds; C486057:dimethyl thiophosphite; D000249:Adenosine Monophosphate; D015032:Zinc