Introduction of chemically labile substructures into Arabidopsis lignin through the use of LigD, the C-dehydrogenase from Sphingobium sp strain SYK-6

• Main Authors: Tsuji, Yukiko, Vanholme, Ruben, Tobimatsu, Yuki, Ishikawa, Yasuyuki, Foster, Clifton E., Kamimura, Naofumi, Hishiyama, Shojiro, Hashimoto, Saki, Shino, Amiu, Hara, Hirofumi, Kanna, Sato-Izawa, Oyarce, Paula, Goeminne, Geert, Morreel, Kris, Kikuchi Morreel, Jun, Takano, Toshiyuki, Fukuda, Masao, Katayama, Yoshihiro, Boerjan, Wout, John, Ralph, Masai, Eiji, Kajita, Shinya
• Format: Article
• Published: Blackwell Publishing 2015
• Subjects: T Technology (General)
• Online Access: http://eprints.utm.my/id/eprint/56015/; http://dx.doi.org/0.1111/pbi.12316

Bacteria-derived enzymes that can modify specific lignin substructures are potential targets to engineer plants for better biomass processability. The Gram-negative bacterium Sphingobium sp. SYK-6 possesses a Ca-dehydrogenase (LigD) enzyme that has been shown to oxidize the a-hydroxy functionalities in ß-O-4-linked dimers into a-keto analogues that are more chemically labile. Here, we show that recombinant LigD can oxidize an even wider range of ß-O-4-linked dimers and oligomers, including the genuine dilignols, guaiacylglycerol-ß-coniferyl alcohol ether and syringylglycerol-ß-sinapyl alcohol ether. We explored the possibility of using LigD for biosynthetically engineering lignin by expressing the codon-optimized ligD gene in Arabidopsis thaliana. The ligD cDNA, with or without a signal peptide for apoplast targeting, has been successfully expressed, and LigD activity could be detected in the extracts of the transgenic plants. UPLC-MS/MS-based metabolite profiling indicated that levels of oxidized guaiacyl (G) ß-O-4-coupled dilignols and analogues were significantly elevated in the LigD transgenic plants regardless of the signal peptide attachment to LigD. In parallel, 2D NMR analysis revealed a 2.1- to 2.8-fold increased level of G-type a-keto-ß-O-4 linkages in cellulolytic enzyme lignins isolated from the stem cell walls of the LigD transgenic plants, indicating that the transformation was capable of altering lignin structure in the desired manner