Reducing lignin levels can lead to lower recalcitrance and higher saccharification efficiency for bio-fuel purposes. Manipulating the monolignol biosynthesis pathway has been the main approach to reducing lignin levels in plants. All enzymes necessary for the formation of the three monolignol building blocks have been characterized, mostly in dicotyledonous plants. We have been characterizing the genes encoding monolignol biosynthesis enzymes in switchgrass.
Our core strategy has been to identify cDNAs in switchgrass databases similar to previously characterized cDNAs from other species; to perform phylogenetic analysis of the translated cDNAs; to heterologously express the genes in E. coli; to characterize the corresponding enzymes biochemically; and to analyze their gene expression patterns in the plant by qRT-PCR.
To date, our targets have been a) 4CL (4-coumarate coenzyme A ligase), b) HCT (hydroxycinnamoyl-CoA:shikimate/quinate hydroxycinnamoyltransferase), c) CCR (cinnamoyl coenzyme A reductase) and d) CAD (cinnamyl alcohol dehydrogenase). The expressed proteins of all of these candidates have shown the predicted biochemical activities and their gene expression follows the pattern seen for genes involved in lignification.
Comparison of the enzyme kinetics of the switchgrass enzymes to those of other species will be discussed, and we will also present an overview of the monolignol pathway in switchgrass.