Arabidopsis CYP707As encode (+)-abscisic acid 8'-hydroxylase, a key enzyme in the oxidative catabolism of abscisic acid.
Abscisic acid (ABA) is involved in a number of critical processes in normal growth and development as well as in adaptive responses to environmental stresses. For correct and accurate actions, a physiologically active ABA level is controlled through fine-tuning of de novo biosynthesis and catabolism. The hydroxylation at the 8'-position of ABA is known as the key step of ABA catabolism, and this reaction is catalyzed by ABA 8'-hydroxylase, a cytochrome P450. Here, we demonstrate CYP707As as the P450 responsible for the 8'-hydroxylation of (+)-ABA. First, all four CYP707A cDNAs were cloned from Arabidopsis and used for the production of the recombinant proteins in insect cells using a baculovirus system. The insect cells expressing CYP707A3 efficiently metabolized (+)-ABA to yield phaseic acid, the isomerized form of 8'-hydroxy-ABA. The microsomes from the insect cells exhibited very strong activity of 8'-hydroxylation of (+)-ABA (K(m) = 1.3 microm and k(cat) = 15 min(-1)). The solubilized CYP707A3 protein bound (+)-ABA with the binding constant K(s) = 3.5 microm, but did not bind (-)-ABA. Detailed analyses of the reaction products confirmed that CYP707A3 does not have the isomerization activity of 8'-hydroxy-ABA to phaseic acid. Further experiments revealed that Arabidopsis CYP707A1 and CYP707A4 also encode ABA 8'-hydroxylase. The transcripts of the CYP707A genes increased in response to salt, osmotic, and dehydration stresses as well as ABA. These results establish that the CYP707A family plays a key role in regulating the ABA level through the 8'-hydroxylation of (+)-ABA.
Saito
Shigeki
S
Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan.
Hirai
Nobuhiro
N
Matsumoto
Chiaki
C
Ohigashi
Hajime
H
Ohta
Daisaku
D
Sakata
Kanzo
K
Mizutani
Masaharu
M
D000040:Abscisic Acid / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000187:drug effects / Q000235:genetics; D060406:Brassinosteroids; D002777:Cholestanols / Q000494:pharmacology; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D005875:Gibberellins / Q000494:pharmacology; D007313:Insecta / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008861:Microsomes / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D010084:Oxidation-Reduction / Q000187:drug effects; D010802:Phylogeny; D010937:Plant Growth Regulators / Q000494:pharmacology; D010940:Plant Proteins; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D013260:Steroids, Heterocyclic / Q000494:pharmacology
D029681:Arabidopsis Proteins; D060406:Brassinosteroids; D002777:Cholestanols; D018076:DNA, Complementary; D005875:Gibberellins; D007527:Isoenzymes; D010937:Plant Growth Regulators; D010940:Plant Proteins; D013260:Steroids, Heterocyclic; D000040:Abscisic Acid; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; C107203:abscisic acid 8'-hydroxylase; C023623:brassinolide