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This presentation discusses aims to: "Communicate the excitement of big science"; Advertise the LANL ICF/HED program; and, Provide a strategic program view for the work and how we use the data.
The work performed at Clark University (PI, David Hibbett) was a subcomponent of the research lead by Jonathan Schilling at University of Minnesota (award number DE-SC0019427). The overall goal of this project was to address how fungi metabolize lignocellulose. Specifically, we targeted a carbohydrate-selective group of fungi known as 'brown rot' fungi that are well-aligned to be harness for bioconversion to biofuels and other products. These organisms orchestrate a two-step mechanism that initiates decomposition using oxidative radical-based chemistry and follows with enzymatic attack. We have helped characterize that this mechanism occurs in this sequence, but currently lack the fundamental regulatory information needed to harness these fungi for application. Thus, the major goal of this project was to use systems biology to create an integrated model of brown rot wood decay that couples fungal gene regulation with the metabolite feedbacks used to coordinate a two-step oxidative-hydrolytic mechanism. The research at Clark University focused on phylogenomic analyses of data produced by other project partners. Hence, the Clark component of the research was staged to initiate somewhat after that in other partner laboratories. The major Clark-based accomplishment was to advise on selection of fungal species for analyses of brown rot decay chemistry, and to interpret results in a phylogenetic context. The Clark University group also performed complementary analyses of gene families encoding plant cell wall-degrading enzymes in the the basidiomycete genus Lentinula, as well as analyses of substrate-specific gene expression and RNA editing in brown rot fungi.
The goal of this Case Study was to quantify the impacts of variable moisture and ash on hammer mill throughput and energy consumption and on generation of fines that are not able to be fed to conversion, as compared to a status quo Base Case system. Also considered was convertible carbon content (minimum carbon specification) and maximum ash content and the delivered feedstock cost impacts of not being able to feed residue not meeting both specifications to the conversion reactor. Laboratory data on the impacts of input particle size and moisture content on the exit particle size were received from FCIC Subtask 5.2 from their single particle impact population balance modeling study. Additional throughput and energy consumption data were obtained from FCIC Subtask 5.2 for the same grinder with a 6 mm screen in place. These data were utilized to develop the necessary response surface equations to perform throughput analysis using discrete event simulation. Because the ash contents in the separated fines had not been analyzed in the laboratory at the time of the model runs, we chose to assume that the ash distributed proportionally with total mass into the overs and unders in the disk screen following grinding. Key takeaways from this Case Study are that it is significantly more cost effective to hammer mill the residue prior to drying, even though the grinder throughput is lower and energy consumption is higher versus drying first before grinding. An effect of dry grinding versus high moisture grinding is the production of higher amounts of fines during dry grinding, leading to significantly more of the ground feedstock being rejected by conversion for being below a minimum particle size. With wet grinding the system is still able to produce more preprocessed feedstock meeting the minimum particle size specification even though the instantaneous throughput is lower than for the case of grinding dry feedstock. Additionally, even without the higher fines production from dry grinding, the status quo would still be more costly than wet grinding because the material is rejected after the drying energy has already been input for the dry grinding case. Finally, significant reductions in drying energy are obtained by drying after grinding, and those reductions are of far greater magnitude than the grinding energy increase.
The goal of this project is to build a multimodal optical microscope to measure the binding, processive degradation, and pausing behaviors of cellulases as they interact with and degrade both synthetic and naturally occurring lignocellulosic walls. To achieve this, we will use high spatio-temporal single-molecule imaging to track cellulases, while visualizing specific molecular components of cellulose, lignin and hemicellulose, that make up their lignocellulose substrate. The microscope will combine Interferometric Scattering (iSCAT), which provides unprecedented spatiotemporal resolution; Total Internal Reflection Fluorescence (TIRF), which provides single-molecule resolution of multiple fluorophore-labeled molecules; and Stochastic Reconstruction (STORM), which allows for three-dimensional super-resolution imaging of intact plant cell walls during degradation. Initial studies will investigate cellulase dynamics on in vitro-assembled cell wall analogs, and later work will progress to using native plant cell walls.
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The National Renewable Energy Laboratory (NREL) developed low severity Deacetylation and Mechanical Refining (DMR) process under atmospheric pressure, generates highly fermentable, low toxicity sugar syrups and highly reactive lignin streams. The dilute alkali deacetylation step saponifies acetyl groups from the hemicellulosic fraction of biomass into a black liquor waste stream that also contains solubilized acetic, ferulic, p-coumaric acids, lignin-carbohydrate complexes (LCCs), oligosaccharides, and solubilized lignin. Valorization of the soluble components in the deacetylation black liquor waste stream requires characterization of the black liquor. Our analyses show that glucan, xylan, and lignin were three main components in black liquor and the oligosaccharides in the black liquor were mainly dimers and trimers with arabinofuranose branch groups mainly on the xylooligomers. GPC chromatograms showed that the black liquor also contained oligomeric lignin moieties with molecular weights ranging between 1,800 and ~10,000 Da. Enzymatic saccharification of the black liquor was conducted based on the compositional and structural information. Incorporation of commercial cellulase and hemicellulase enzyme preparations with various accessary enzymes showed up to 57% hydrolysis of the xylooligosaccharides in the black liquor to monomeric sugars, which could be used to improve biorefinery carbon utilization in biological fermentations to improve end product fuel or bioproduct volumes per metric ton of biomass processed. This is so far the first study about characterizing DMR black liquor and exploring breaking down the oligosaccharides in black liquor for potential use.