Poster: Cell Walls

Abs # 1251: Cinnamoyl-Coenzyme A Reductase Reaction Profiles Suggest A Main Metabolic Flux for Monolignol Biosynthesis in Woody Xylem

Presenter:	Li, Laigeng , Laigeng_Li@ncsu.edu
Authors	Li, Laigeng  (A)   Cheng, Xiaofei  (B)   Lu, Shanfa  (A)   Umezawa, Toshiaki  (C)   Chiang, Vincent L. (A)
Affiliations:	(A): North Carolina State University (B): FBG, Noble Foundation (C): Kyoto University

Cinnamoyl-Coenzyme A Reductase (CCR), one of the key enzymes involved in the biosynthesis of monolignols, has been thought to catalyze the conversion of various cinnamoyl CoA esters to their corresponding cinnamaldehydes. This had lead to the suggestion of a metabolic grid for monolignol biosynthesis. A xylem-specific CCR cDNA was cloned from aspen developing xylem tissue. The recombinant CCR protein was produced through an E. coli expression system and purified to electrophoretic homogeneity. CCR biochemical properties were thoroughly characterized through direct structural corroboration and quantitation of the reaction products using a liquid chromatography-mass spectrometry system. The enzyme kinetics profiling demonstrated that CCR was highly specific for catalyzing the reduction of feruoyl-CoA as measured by Km and Vmax/Km values, and that when various cinnamoyl CoA esters were used as a mixed substrate pool, feruoyl-CoA became a strong competitive inhibitor for CCR reactions with other CoA esters. Moreover, when caffeoyl-CoA O-methyltransferase (CCoAOMT) was coupled with CCR in a reaction with caffeoyl-CoA ester as the substrate, caffeoyl-CoA was converted into coniferaldehyde but not caffealdehyde. Taken together, the results suggested that the CCoAOMT/CCR-mediated conversion of caffeoyl-CoA ester to coniferaldehyde via feruoyl-CoA constituted a main metabolic flux toward monolignol biosynthesis in lignifying xylem tissue.