title: Purification and characterization of isoforms of cinnamyl alcohol dehydrogenase from Eucalyptus xylem.
abstract: Two distinct isoforms of cinnamyl alcohol dehydrogenase, CAD 1 and CAD 2, have been purified to homogeneity from xylem-enriched fractions of Eucalyptus gunii Hook and partially characterized. They differ greatly in terms of both physical and biochemical properties, and can be separated by hydrophobic interaction chromatography on Phenyl Sepharose CL-4B. The native molecular weight of of CAD 1 is 38 kDa as determined by gel-filtration chromatography on Superose 6, and this isoform is likely to be a monomer since it yields a polypeptide of 35 kDa upon sodium dodecyl sulfatepolyacrylamide gel electrophoresis. It has a low substrate affinity for coniferyl and p-coumaryl alcohols and their corresponding aldehydes. No activity with sinapyl aldehyde and alcohol was detected. The more abundant isoform is CAD 2, which has a native molecular weight of 83 kDa and is a dinier composed of two subunits of slightly different molecular weights (42-43 kDa). These subunits show identical peptide patterns after digestion with N-chlorosuccinimide. The isoform, CAD 2, has a high substrate affinity for all the substrates tested. The two isoforms are immunologically distinct as polyclonal antibodies raised against CAD 2 do not cross-react with CAD 1. The characterization of two forms of CAD exhibiting such marked differences indicates their involvement in specific pathways of monolignol utilisation.
authors.affiliation: Université Paul Sabatier, Centre de Biologie et Physiologie Végétale, URA CNRS 1457, 118, route de Narbonne, F-31062, Toulouse Cedex, France.
authors.forename: D
authors.initials: D
authors.lastname: Goffner
authors.forename: I
authors.initials: I
authors.lastname: Joffroy
authors.forename: J
authors.initials: J
authors.lastname: Grima-Pettenati
authors.forename: C
authors.initials: C
authors.lastname: Halpin
authors.forename: M E
authors.initials: ME
authors.lastname: Knight
authors.forename: W
authors.initials: W
authors.lastname: Schuch
authors.forename: A M
authors.initials: AM
authors.lastname: Boudet