Microsomally associated heme proteins from French bean: characterization of the cytochrome P450 cinnamate-4-hydroxylase and two peroxidases.

Three heme-containing proteins of M(r) 57,000, 46,000, and 42,000 have been isolated from microsomes of elicitor-treated suspension-cultured cells of French bean. Following purification to homogeneity they were shown to be distinct with respect to amino acid and sugar compositions. The M(r) 57,000 protein proved to be the cytochrome P450 cinnamate-4-hydroxylase (CYP 73), by immunopurification and characterization. The M(r) 46,000 showed peroxidative activity toward guaiacol and all substituted phenylpropanoids tested and also gave a form of binding spectrum with cinnamic acid. The M(r) 42,000 had peroxidative activity toward guaiacol and a similar apparent binding spectrum to the M(r) 46,000 protein but was less active toward hydroxylated and methoxylated phenylpropanoids. The two peroxidases had cationic pIs and were distinct proteins rather than glycosylated or proteolytic variants.