Uncovering the Physico-Chemical Rules Of Assembly in Grass Cell Wall Architecture that Lead to Enzymatic Recalcitrance via Multi-Dimensional Solid State Nuclear Magnetic Resonance
EMSL Project ID
48883
Abstract
Advanced solid state (ss) NMR methods will be used to elucidate the architecture and arrangement of grass plant cell walls. Our understanding of physical interactions in intact cell walls between cellulose and hemicellulose components, is very limited. These interactions contribute to cell wall cohesiveness and recalcitrance to enzymatic deconstruction. Previous ssNMR work has investigated model systems, such as Arabidopsis and Brachypodium, whereas most cellulosic feedstocks in the future are expected to come from sugarcane, switchgrass, Miscanthus, maize and other species in the grass family (Poaceae). Our proposed study will make use of state-of-the-art multi-dimensional ssNMR methods and 13C-enriched cell walls from the grass species Zea mays (=maize, corn) to address a specific interaction between beta(1-3, 1-4)-D-glucan (not present in Arabidopsis or Brachypodium) and cellulose microfibrils. This proposal concentrates on several specific questions regarding the architecture of grass cell walls: (a) What proportion of cellulose surfaces are bound to hemicelluloses? (b) What is the proportion of surface bound to beta(1-3, 1-4)-D-glucan, arabinoxylan, and xyloglucan (these are the dominant hemicelluloses in the typical primary cell wall of grasses, in order of abundance)? (c) Does the hemicellulose beta(1-3, 1-4)-D-glucan interact closely with the cellulose surface? (d) Are hemicelluloses bound or entangled with each other, at some distance from the cellulose surface? (e) Do pectins play a larger role than previously predicted in grass cell walls? (f) What fraction of hemicellulosic polymers is mobile, and what fraction is rigid (thus presumably bound to cellulose surfaces)? (g) Are polymer interactions within the grass cell wall consistent with current hypotheses of cell wall structure? This knowledge is relevant to the EMSL science theme entitled Biosystem Dynamics and Design because characterization of the nano-scale grass cell wall components will reveal the complex polymer network of plant construction. In the first year of these proposed studies, we will collect 2D ssNMR to further elucidate structural differences of grass cell wall, using 13C-labeled Z. mays cell wall samples at both 400 and 850 MHz field strengths. 13C spin-lattice relaxation data will also be acquired at this time, using different excitation methods to distinguish between mobile and rigid components, with an emphasis on dynamics. Additional multi-dimensional experiments, as described in the proposal, will uncover new structural information about the hemicellulosic polymers and their interactions, specifically beta(1-3, 1-4)-D-glucans. Our results will explicate the key physical interactions underlying the cell wall architecture of grass species, thereby advancing the plant cell wall community significantly. This study was conceived within and will be carried out with strong participation from the DOE-funded Energy Frontier Research Center (EFRC) on Lignocellulose Structure and Formation (CLSF), directed by the PI of this proposal. The NMR capabilities and expertise at EMSL are critical for this proposal: the resolution and sensitivity enhancements obtained from running experiments at 20.0 T will provide unprecedented molecular-scale details for interfacial components in this important system.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2015-10-01
End Date
2017-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members