ARS | Publication request: Fluorescence-tagged monolignols: Synthesis, and application to studying in vitro lignification

Submitted to: Biomacromolecules
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 16, 2011
Publication Date: March 31, 2011
Citation: Tobimatsu, Y., Davidson, C.L., Grabber, J.H., Ralph, J. 2011. Fluorescence-tagged monolignols: Synthesis, and application to studying in vitro lignification. Biomacromolecules. 12: 1752-1761.

Interpretive Summary: Plants are mainly composed of cell walls, which are oftentimes referred to as fiber. Plant cell walls are primarily composed of polysaccharides (polymers of sugar molecules) cemented together with another polymer known as lignin. Although lignin plays a number of important roles in plants, its presence severely limits the utilization of cell wall polysaccharides as a source of digestible energy for livestock and as a source of sugars for the industrial production of liquid fuels and other products. The removal of lignin from fiber also greatly increases the cost and environmental impact of converting woody plants into paper products. Thus scientists have long studied the biosynthesis of monolignols (lignin building blocks) and their polymerization into lignin with the aim of manipulating the process to lessen the inhibitory effects of lignin on fiber utilization. In this study, we developed methods for attaching florescent tags to monolignols and demonstrated in the laboratory that the modified monolignols readily diffused into cell walls, interacted with peroxidase (an enzyme responsible for lignin polymerization), and formed lignin polymers. We anticipate fluorescence-tagged monolignols might be useful for visualizing the transport and polymerization of monolignols in plants. Such studes may reveal new ways of altering lignification process to improve plant cell wall utilization by livestock and industry.

Technical Abstract: Coniferyl alcohol gamma-coupled by ethylenediamine spacers to dimethylaminocoumarin or nitrobenzofuran fluorophores was tested as a photoprobe to study the oxidase-mediated polymerization of monolignols. The fluorescent coniferyl alcohol derivatives readily underwent peroxidase-catalyzed in vitro copolymerization with coniferyl alcohol to yield fluorescent dehydrogenation polymers that were structurally indistinguishable from polymers formed solely from coniferyl alcohol. To illustrate the use of the photoprobes, we successfully monitored in real time the complexation of coniferyl alcohol with horseradish apoperoxidase by Förster resonance energy transfer (FRET), using the protein-tryptophan and a dimethylaminocoumarin moiety as donor and acceptor fluorophores. Furthermore mixtures of fluorescent-tagged and normal coniferyl alcohols readily diffused into isolated maize cell walls and reacted with wall-bound peroxidases to form artificial lignins that could be visualized by fluorescent microscopy. Thus we anticipate fluorescence-tagged monolignols will be useful for in vitro and in vivo studies of cell wall lignification.