Structures of Michaelis and product complexes of plant cytokinin dehydrogenase: Implications for flavoenzyme catalysis

被引:68
作者
Malito, E
Coda, A
Bilyeu, KD
Fraaije, MW
Mattevi, A
机构
[1] Univ Pavia, Dept Genet & Microbiol, I-27100 Pavia, Italy
[2] Univ Missouri, USDA ARS, Columbia, MO 65211 USA
[3] Univ Groningen, Biochem Lab, NL-9747 AG Groningen, Netherlands
关键词
flavoenzyme; mechanism of catalysis; cytokinin; crystallography;
D O I
10.1016/j.jmb.2004.06.083
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Cytokinins form a diverse class of compounds that are essential for plant growth. Cytokinin dehydrogenase has a major role in the control of the levels of these plant hormones by catalysing their irreversible oxidation. The crystal structure of Zea mays cytokinin dehydrogenase displays the same two-domain topology of the flavoenzymes of the vanillyl-alcohol, oxidase family but its active site cannot be related to that of any other family member. The X-ray analysis reveals a bipartite architecture of the, catalytic centre, which consists of a funnel-shaped region on the protein surface and an internal cavity lined by the flavin ring. A pore with diameter of about 4 A connects the two active-site regions. Snapshots of two critical steps along the reaction cycle were obtained through the structural analysis of the complexes with a slowly reacting substrate and the reaction product, which correspond to the states immediately before (Michaelis complex) and after (product complex) oxidation has taken place. The substrate displays a "plug-into-socket" binding mode that seals the catalytic site and precisely positions the carbon atom undergoing oxidation in close contact with the reactive locus of the flavin. A polarising H-bond between the substrate amine group and an Asp-Glu pair may facilitate oxidation. Substrate to product conversion results in small atomic movements, which lead to a planar conformation of the reaction product allowing double-bond conjugation. These features in the mechanism of amine recognition and oxidation differ from those observed in other flavin-dependent amine oxidases. (C) 2004 Elsevier Ltd. All rights reserved.
引用
收藏
页码:1237 / 1249
页数:13
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