ISOLATION AND SEQUENCE OF A CDNA-ENCODING THE JERUSALEM-ARTICHOKE CINNAMATE 4-HYDROXYLASE, A MAJOR PLANT CYTOCHROME-P450 INVOLVED IN THE GENERAL PHENYLPROPANOID PATHWAY
Hg. Teutsch et al., ISOLATION AND SEQUENCE OF A CDNA-ENCODING THE JERUSALEM-ARTICHOKE CINNAMATE 4-HYDROXYLASE, A MAJOR PLANT CYTOCHROME-P450 INVOLVED IN THE GENERAL PHENYLPROPANOID PATHWAY, Proceedings of the National Academy of Sciences of the United Statesof America, 90(9), 1993, pp. 4102-4106
Cinnamate 4-hydroxylase [CA4H; trans-cinnamate, NADPH:oxygen oxidoredu
ctase (4-hydroxylating), EC 1.14.13.11] is a cytochrome P450 that cata
lyzes the first oxygenation step of the general phenylpropanoid metabo
lism in higher plants. The compounds formed are essential for lignific
ation and defense against predators and pathogens. We recently reporte
d the purification of this enzyme from Mn2+-induced Jerusalem artichok
e (Helianthus tuberosus L.) tuber tissues. Highly selective polyclonal
antibodies raised against the purified protein were used to screen a
lambdagt11 cDNA expression library from wound-induced Jerusalem artich
oke, allowing isolation of a 1130 -base-pair insert. Typical P450 doma
ins were identified in this incomplete sequence, which was used as a p
robe for the isolation of a 1.7-kilobase clone in a lambdagt10 library
. A full-length open reading frame of 1515 base pairs, encoding a P450
protein of 505 residues (M(r) = 57,927), was sequenced. The N terminu
s, essentially composed of hydrophobic residues, matches perfectly the
microsequenced N terminus of the purified protein. The calculated pI
is 9.78, in agreement with the chromatographic behavior and two-dimens
ional electrophoretic analysis of CA4H. Synthesis of the corresponding
mRNA is induced in wounded plant tissues, in correlation with CA4H en
zymatic activity. This P450 protein exhibits the most similarity (28%
amino acid identity) with avocado CYP71, but also good similarity with
CYP17 and CYP21, or with CYP1 and CYP2 families. According to current
criteria, it qualifies as a member of a new P450 family.