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DOE OSTI2020

Life cycle Management of 324 Project High-Activity, Mixed/Low-Level Radioactive Waste - 20506

U.S. DOE contractor CH2M Hill Plateau Remediation Company (CHPRC) manages the 324 Building and is preparing to remotely excavate and disposition the radioactive soil beneath the building to allow further deactivation of the building. The 324 Building is a non-reactor Category 2 Nuclear Facility located in the 300 Area of the Hanford Site. Records indicate that in October 1986, approximately 516 liters of a concentrated liquid waste stream containing cesium-137 ({sup 137}Cs) and strontium-90 ({sup 90}Sr) spilled onto the floor of a hot cell, B-Cell, in the 324 Building. The spill contained an estimated 1.3 million curies (Ci) of radioactivity. A breach in the sump of B-Cell was discovered in November 2009 during characterization of the soil under the building. Exposure rates in excess of 10,000 R/hr were detected through a system of access pipes installed under the hot cell, and an estimated 1.557 E+05 Ci of {sup 137}Cs and 6.842 E+04 Ci of {sup 90}Sr are in the soil beneath B-Cell. The magnitude of the soil contamination below B-Cell represents one of the most challenging remediation activities in the DOE complex. The objective of CHPRC's 324 Project is to remotely excavate and disposition the highly contaminated soil under the hot cell so building deactivation and stabilization activities can resume. Depending on the volume and radioactivity of contaminated soil under B-Cell, the material will be dispositioned either at the Hanford Site's Environmental Restoration Disposal Facility (ERDF) or in grout monoliths that will be created in the hot cells (A, C and D) next to B-Cell during future building demolition activities. The current project scope includes installing structural supports under the walls of B-Cell and using remotely operated equipment to remove debris from the hot cell, remove debris mixed with grout on the floor, saw through and remove the cell floor and liner, and excavate contaminated soil under B-Cell to a depth of up to 3.66 meters (12 feet). Addition or removal of debris in A-Cell may be required to support B-Cell cleanout and excavation activities. The base approach is to disposition debris and soil bins in the Radiochemical Engineering Cell (REC) monoliths, and subsequently disposition the monoliths during building demolition. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗

DOE OSTI2025

Hemicellulose Modulates Nanoscale Lignin Architecture in Synthetic Plant Cell Walls

Cellulose, hemicellulose, and lignin─the most abundant biopolymers on Earth─compose the structural matrix of plant biomass, providing renewable resources critical to bioenergy and sustainable materials. Despite their importance, the nanoscale mechanochemical processes underlying lignocellulose assembly during plant secondary cell wall formation remain poorly understood, hindering advancements in biomass conversion technologies. Here, in this study, we synthesize a biomimetic model system comprising cellulose–hemicellulose nanofibrils (CHN) to examine guaiacyl lignin polymerization in a physiologically relevant context. Using advanced nanocharacterization─scattering-type scanning near-field optical microscopy (s-SNOM) with infrared nanospectroscopy coupled to solid-state nuclear magnetic resonance (NMR)─we reveal that hemicellulose presence considerably modulates lignin deposition and alters its interunit bond distribution. Specifically, hemicellulose-rich environments dramatically reduce lignin deposition by approximately 50% and yield highly condensed lignin structures characterized by severely reduced β–O–4′ linkages (<2%) and suppressed β–β′ linkages. Conversely, cellulose-alone scaffolds support notably higher β–O–4′ content (∼10%), resulting in a more uniform nanoscale lignin coating. Our work helps explain how accessible hemicellulose sites, both sterically and chemically, direct radical coupling during lignification, fundamentally reshaping lignin’s nanoscale architecture. These findings deepen our mechanistic understanding of plant cell wall biosynthesis and inform strategies aimed at enhancing biomass deconstruction efficiency for sustainable bioenergy applications.

NMR↗

DOE OSTI2026

Native Architecture of Wheat Straw Cell Walls: A Unified Model from X-ray Scattering and Solid-State NMR

Plant secondary cell walls constitute the dominant reservoir of renewable biomass, comprising tightly packed cellulose, hemicellulose, and lignin at the nanoscale. Recent advances in solid-state NMR spectroscopy and the availability of small-angle X-ray scattering for biomass characterization have led to an accumulation of experimental data on cell wall organization, yet no explicit structure model has simultaneously satisfied both Xray and NMR observations. Using wheat straw as a model system, we propose a structural framework consistent with current knowledge of cellulose biosynthesis, X-ray scattering data, and one- and two-dimensional 13 C solid-state NMR spectra. In this model, 18-chain elementary fibrils align in parallel and populate the cross-section at random. Arabinose-substituted xylan shows no conformational dependence for cellulose-binding in wheat, and only a minor fraction of 2-fold xylan appears in close proximity to cellulose, unlike in Arabidopsis, where xylan is more tightly attached to the cellulose surface. While NMR data cannot unambiguously resolve the internal arrangement of the 18 glucan chains, X-ray scattering profiles uniquely constrain the fibril size and exclude the possibility of tight bundling in the intact walls. The specific interaction between the matrix polymers and the cellulose elementary fibrils must be reconsidered in light of the small interfibril spaces, which bring the matrix components into spatial proximity with cellulose even in the absence of attractive interactions. These findings provide fundamental molecular-level insight into cellulose fibril architecture and matrix−polymer interactions, resolving longstanding discrepancies between spectroscopic and scattering data and advancing our understanding of biopolymer assembly into structurally and functionally versatile lignocellulosic biomaterials.

Carbohydrates↗

NASA NTRS1989

Microsandwich cellular solids

This article introduces a new concept of microsandwiching in which the cell wall itself is a sandwich structure and is substantially stiffer. Some experimental results are presented to show the efficacy of this approach. A method for optimizing the design of honeycomb microsandwich is also briefly outlined.

Balakrishna Bhat, T.↗

NASA NTRS1999

In vitro activities of four xyloglucan endotransglycosylases from Arabidopsis

Xyloglucan endotransglycosylases (XETs) are encoded by a gene family in Arabidopsis thaliana. These enzymes modify a major structural component of the plant cell wall, xyloglucan, and therefore may influence plant growth and development. We have produced four Arabidopsis XETs (TCH4, Meri-5, EXGT and XTR9) using the baculovirus/insect cell system and compared their biochemical activities. TCH4, as previously demonstrated, and the other three proteins are capable of carrying out transglycosylation of xyloglucans. The K(m) for XLLGol acceptor oligosaccharide is in the range of 20-40 microM for all the XETs except XTR9, which has a Km of 5 microM and is significantly inhibited by high levels of XLLGol. All four enzymes are most active between pH 6.0 and 6.5. TCH4 and XTR9 have temperature optima of 18 degrees C, whereas Meri-5 and EXGT are most active at 28 and 37 degrees C, respectively. Although the activity levels of three of the XETs are not influenced by the presence of fucose on the xyloglucan polymer, XTR9 has a clear preference for non-fucosylated xyloglucan polymer. The four XETs show a marked preference for XLLGol over either XXFGol or XXXGol as acceptor oligosaccharide. All four XETs are glycosylated; however, only the activities of TCH4 and Meri-5 are affected by the removal of the N-glycan with PNGase F. These four enzymes most likely function solely as transglycosylases because xyloglucan endoglucanase activity was not apparent. Subtle differences in biochemical activities may influence the physiological functions of the distinct XETs in vivo.

Non-NASA Center↗

DOE OSTI2021

A pectin methyltransferase modulates polysaccharide dynamics and interactions in Arabidopsis primary cell walls: Evidence from solid-state NMR

Plant cell walls contain cellulose embedded in matrix polysaccharides. Understanding carbohydrate structures and interactions is critical to the production of biofuel and biomaterials using these natural resources. Here we present a solid-state NMR study of cellulose and pectin in 13 C-labeled cell walls of Arabidopsis wild-type and mutant plants. Using 1D 13 C and 2D 13 C– 13 C correlation experiments, we detected a highly branched arabinan structure in qua2 and tsd2 samples, two allelic mutants for a pectin methyltransferase. Both mutants show close physical association between cellulose and the backbones of pectic homogalacturonan and rhamnogalacturonan-I. Relaxation and dipolar order parameters revealed enhanced microsecond dynamics due to polymer disorder in the mutants, but restricted motional amplitudes due to tighter pectin-cellulose associations. These molecular data shed light on polymer structure and packing in these two pectin mutants, helping to elucidate how pectin could influence cell wall architecture at the nanoscale, cell wall mechanics, and plant growth.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

DOE OSTI2022

Current limitations of solid-state NMR in carbohydrate and cell wall research

High-resolution investigation of cell wall materials has emerged as an important application of biomolecular solid-state NMR (ssNMR). Multidimensional correlation experiments have become a standard method for obtaining sufficient spectral resolution to determine the polymorphic structure of carbohydrates and address biochemical questions regarding the supramolecular organization of cell walls. Here, using plant cellulose and matrix polysaccharides as examples, we will review how the multifaceted complexity of polysaccharide structure is impeding the resonance assignment process and assess the available biochemical and spectroscopic approaches that could circumvent this barrier. We will emphasize the ineffectiveness of the current methods in reconciling the ever-growing dataset and deriving structural information. We will evaluate the protocols for achieving efficient and homogeneous hyperpolarization across the cell wall material using magic-angle spinning dynamic nuclear polarization (MAS-DNP). Critical questions regarding the line-broadening effects of cell wall molecules at cryogenic temperature and by paramagnetic biradicals will be considered. Finally, the MAS-DNP method will be placed into a broader context with other structural characterization techniques, such as cryo-electron microscopy, to advance ssNMR research in carbohydrate and cell wall biomaterials.

09 BIOMASS FUELS↗

NASA NTRS2001

Dynamic Finite Element Predictions for Mars Sample Return Cellular Impact Test #4

The nonlinear finite element program MSC.Dytran was used to predict the impact pulse for (he drop test of an energy absorbing cellular structure. This pre-test simulation was performed to aid in the design of an energy absorbing concept for a highly reliable passive Earth Entry Vehicle (EEV) that will directly impact the Earth without a parachute. In addition, a goal of the simulation was to bound the acceleration pulse produced and delivered to the simulated space cargo container. EEV's are designed to return materials from asteroids, comets, or planets for laboratory analysis on Earth. The EEV concept uses an energy absorbing cellular structure designed to contain and limit the acceleration of space exploration samples during Earth impact. The spherical shaped cellular structure is composed of solid hexagonal and pentagonal foam-filled cells with hybrid graphite-epoxy/Kevlar cell walls. Space samples fit inside a smaller sphere at the enter of the EEV's cellular structure. The material models and failure criteria were varied to determine their effect on the resulting acceleration pulse. Pre-test analytical predictions using MSC.Dytran were compared with the test results obtained from impact test #4 using bungee accelerator located at the NASA Langley Research Center Impact Dynamics Research Facility. The material model used to represent the foam and the proper failure criteria for the cell walls were critical in predicting the impact loads of the cellular structure. It was determined that a FOAMI model for the foam and a 20% failure strain criteria for the cell walls gave an accurate prediction of the acceleration pulse for drop test #4.

Fasanella, Edwin L.↗

DOE OSTI2024

Insights into the action of phylogenetically diverse microbial expansins on the structure of cellulose microfibrils

Microbial expansins (EXLXs) are non-lytic proteins homologous to plant expansins involved in plant cell wall formation. Due to their non-lytic cell wall loosening properties and potential to disaggregate cellulosic structures, there is considerable interest in exploring the ability of microbial expansins (EXLX) to assist the processing of cellulosic biomass for broader biotechnological applications. Herein, EXLXs with different modular structure and from diverse phylogenetic origin were compared in terms of ability to bind cellulosic, xylosic, and chitinous substrates, to structurally modify cellulosic fibrils, and to boost enzymatic deconstruction of hardwood pulp. Five heterogeneously produced EXLXs (Clavibacter michiganensis; CmiEXLX2, Dickeya aquatica; DaqEXLX1, Xanthomonas sacchari; XsaEXLX1, Nothophytophthora sp.; NspEXLX1 and Phytophthora cactorum; PcaEXLX1) were shown to bind xylan and hardwood pulp at pH 5.5 and CmiEXLX2 (harboring a family-2 carbohydrate-binding module) also bound well to crystalline cellulose. Small-angle X-ray scattering revealed a 20–25% increase in interfibrillar distance between neighboring cellulose microfibrils following treatment with CmiEXLX2, DaqEXLX1, or NspEXLX1. Correspondingly, combining xylanase with CmiEXLX2 and DaqEXLX1 increased product yield from hardwood pulp by ~ 25%, while supplementing the TrAA9A LPMO from Trichoderma reesei with CmiEXLX2, DaqEXLX1, and NspEXLX1 increased total product yield by over 35%. This direct comparison of diverse EXLXs revealed consistent impacts on interfibrillar spacing of cellulose microfibers and performance of carbohydrate-active enzymes predicted to act on fiber surfaces. These findings uncover new possibilities to employ EXLXs in the creation of value-added materials from cellulosic biomass.

09 BIOMASS FUELS↗

DOE OSTI2024

The DYRKP1 kinase regulates cell wall degradation in Chlamydomonas by inducing matrix metalloproteinase expression

Abstract The cell wall of plants and algae is an important cell structure that protects cells from changes in the external physical and chemical environment. This extracellular matrix, composed of polysaccharides and glycoproteins, must be constantly remodeled throughout the life cycle. However, compared to matrix polysaccharides, little is known about the mechanisms regulating the formation and degradation of matrix glycoproteins. We report here that a plant kinase belonging to the dual-specificity tyrosine phosphorylation-regulated kinase (DYRKP1) family present in all eukaryotes regulates cell wall degradation after mitosis of Chlamydomonas reinhardtii by inducing the expression of matrix metalloproteinases. Without DYRKP1, daughter cells cannot disassemble parental cell walls and remain trapped inside for more than 10 days. On the other hand, the dual-specificity tyrosine phosphorylation-regulated kinase complementation lines show normal degradation of the parental cell wall. Transcriptomic and proteomic analyses indicate a marked downregulation of MMP gene expression and accumulation, respectively, in the dyrkp1 mutants. The mutants deficient in matrix metalloproteinases retain palmelloid structures for a longer time than the background strain, like dyrkp1 mutants. Our findings show that dual-specificity tyrosine phosphorylation-regulated kinase, by ensuring timely MMP expression, enables the successful execution of the cell cycle. Altogether, this study provides insight into the life cycle regulation in plants and algae.

Kim, Minjae (ORCID:0000000223561295)↗

DOE OSTI2023

Electron tomography unravels new insights into fiber cell wall nanostructure; exploring 3D macromolecular biopolymeric nano-architecture of spruce fiber secondary walls

Lignocellulose biomass has a tremendous potential as renewable biomaterials for fostering the “bio-based society” and circular bioeconomy paradigm. It requires efficient use and breakdown of fiber cell walls containing mainly cellulose, hemicellulose and lignin biopolymers. Despite their great importance, there is an extensive debate on the true structure of fiber walls and knowledge on the macromolecular nano-organization is limited and remains elusive in 3D. We employed dual-axis electron tomography that allows visualization of previously unseen 3D macromolecular organization/biopolymeric nano-architecture of the secondary S2 layer of Norway spruce fiber wall. Unprecedented 3D nano-structural details with novel insights into cellulose microfibrils (~2 nm diameter), macrofibrils, nano-pore network and cell wall chemistry (volume %) across the S2 were explored and quantified including simulation of structure related permeability. Matrix polymer association with cellulose varied between microfibrils and macrofibrils with lignin directly associated with MFs. Simulated bio-nano-mechanical properties revealed stress distribution within the S2 and showed similar properties between the idealized 3D model and the native S2 (actual tomogram). Present work has great potential for significant advancements in lignocellulose research on nano-scale understanding of cell wall assembly/disassembly processes leading to more efficient industrial processes of functionalization, valorization and target modification technologies.

3-D reconstruction↗

DOE OSTI2020

Experimental and Numerical Analysis of Additively Manufactured Inconel 718 Coupons With Lattice Structure

Abstract In the present paper, two lattice geometries suitable for near surface and double wall cooling were developed and tested. The first type of unit cell consisted of six ligaments of 0.5 mm diameter joined at a common vertex near the middle. The second type of unit cell was derived from the first type by adding four mutually perpendicular ligaments in the middle plane. Two lattice configurations, referred to as L1 and L2, respectively, were obtained by repeating the corresponding unit cell in streamwise and spanwise directions in an inline fashion. Test coupons consisting of these lattice geometries embedded inside rectangular cooling channel with dimensions of 2.54 mm height, 38.07 mm width, and 38.1 mm in length were fabricated using Inconel 718 powder and selective laser sintering (SLS) process. The heat transfer and pressure drop performance was then evaluated using steady-state tests with constant wall temperature boundary condition and for channel Reynolds number ranging from 2800 to 15,000. The lattices depicted a higher heat transfer compared with a smooth channel and both the heat transfer and pressure drop increased with a decrease in the porosity from L1 to L2. Steady-state conjugate numerical results revealed formation of prominent vortical structures in the inter-unit cell spaces, which diverted the flow toward the top end wall and created an asymmetric heat transfer between the two end walls. In conclusion, these lattice structures provided an augmented heat transfer while favorably redistributing the coolant within channel.

Engineering↗

DOE OSTI2021

Structures of the mycobacterial membrane protein MmpL3 reveal its mechanism of lipid transport

The mycobacterial membrane protein large 3 (MmpL3) transporter is essential and required for shuttling the lipid trehalose monomycolate (TMM), a precursor of mycolic acid (MA)-containing trehalose dimycolate (TDM) and mycolyl arabinogalactan peptidoglycan (mAGP), in Mycobacterium species, including Mycobacterium tuberculosis and Mycobacterium smegmatis. However, the mechanism that MmpL3 uses to facilitate the transport of fatty acids and lipidic elements to the mycobacterial cell wall remains elusive. Here, we report 7 structures of the M. smegmatis MmpL3 transporter in its unbound state and in complex with trehalose 6-decanoate (T6D) or TMM using single-particle cryo-electron microscopy (cryo-EM) and X-ray crystallography. Combined with calculated results from molecular dynamics (MD) and target MD simulations, we reveal a lipid transport mechanism that involves a coupled movement of the periplasmic domain and transmembrane helices of the MmpL3 transporter that facilitates the shuttling of lipids to the mycobacterial cell wall.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2025

The three cellulose synthase isoforms for secondary cell wall make specific contributions to microfibril synthesis

Cellulose is synthesized at the plasma membrane by the cellulose synthase complex, a structure that contains three distinct isoforms of the catalytic subunit, cellulose synthase A (CESA). The division into three subunits appears early in land plant evolution and is highly conserved, particularly for the secondary cell wall. However, what if any unique roles each isoform plays in the complex remain unclear. Here, we assessed the contributions of specific isoforms to microfibril synthesis. First, we expressed CESA isoforms of the primary cell wall or the moss Physcomitrium patens in Arabidopsis thaliana backgrounds missing a secondary cell wall CESA. While the primary cell wall isoforms rescued the cesa knockout phenotype with partial isoform specificity, those from the moss rescued with fewer restrictions. Then, we recreated various CESA missense mutations in all three of the secondary cell wall isoforms; while results are consistent with isoform specificity, they are difficult to interpret further without molecular structures. Finally, we show that catalytically inactive CESA isoforms restore growth and cellulose content in the corresponding knockout in an isoform-specific manner; along with partial rescue of the growth and cellulose content of the inflorescence stem, the replacement lines have fiber cells with partially disorganized microfibrils and secondary cell wall cellulose with narrow crystal width. Generally, effects were more pronounced in lines where CESA8 was inactivated compared with inactivating CESA4 or 7, which tended to have similar phenotypes to each other. Here, we account for these results with a model for cellulose synthase structure with the isoforms assigned specific localization within the cellulose synthase complex.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Quantification of Native Lignin Structural Features with Gel–Phase 2D–HSQC 0 Reveals Lignin Structural Changes During Extraction

Our ability to study and valorize the lignin fraction of biomass is hampered by the fundamental and still unmet challenge of precisely quantifying native lignin's structural features. Here, we developed a rapid elevated-temperature 1 H– 13 C Heteronuclear Single-Quantum Coherence Zero (HSQC 0 ) NMR method that enables this precise quantification of native lignin structural characteristics even with whole plant cell wall (WPCW) NMR spectroscopy, overcoming fast spin relaxation in the gel phase. We also formulated a Gaussian fitting algorithm to perform automatic and reliable spectral integration. By combining HSQC 0 measurements with yield measurements following depolymerisation, we can confirm the combinatorial nature of radical coupling reactions during biosynthesis leading to a random sequential organization of linkages within a largely linear lignin chain. Such analyses illustrate how this analytical method can greatly facilitate the study of native lignin structure, which can then be used for fundamental studies or to understand lignin depolymerization methods like reductive catalytic fractionation or aldehyde-assisted fractionation.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

DOE OSTI2024

Quantification of Native Lignin Structural Features with Gel–Phase 2D–HSQC 0 Reveals Lignin Structural Changes During Extraction

09 BIOMASS FUELS↗

DOE OSTI2025

Effects of feruloyl-CoA 6'-hydroxylase 1 overexpression on lignin and cell wall characteristics in transgenic hybrid aspen

In plant cell walls, lignin, cellulose, and the hemicelluloses form intricate three-dimensional structures. Owing to its complexity, lignin often acts as a bottleneck for the efficient utilization of polysaccharide components as biochemicals and functional materials. A promising approach to mitigate and/or overcome lignin recalcitrance is the qualitative and quantitative modification of lignin by genetic engineering. Feruloyl-CoA 6'-hydroxylase (F6'H1) is a 2-oxoglutarate-dependent dioxygenase that catalyzes the conversion of feruloyl-CoA, one of the intermediates of the lignin biosynthetic pathway, into 6'-hydroxyferuloyl-CoA, the precursor of scopoletin (7-hydroxy-6-methoxycoumarin). In a previous study with Arabidopsis thaliana, we demonstrated that overexpression of F6'H1 under a xylem-preferential promoter led to scopoletin incorporation into the cell wall. This altered the chemical structure of lignin without affecting lignin content or saccharification efficiency. In the present study, the same F6'H1 construct was introduced into hybrid aspen (Populus tremula × tremuloides T89), a model woody plant, and its effects on plant morphology, lignin chemical structure, global gene expression, and phenolic metabolism were examined. The transgenic plants successfully overproduced scopoletin while exhibiting severe growth retardation, a phenotype not previously observed in Arabidopsis. Scopoletin accumulation was most pronounced in the secondary walls of tracheary elements and the compound middle lamella, with low levels in the fiber cell walls. Overexpression of F6'H1 also affected the metabolism of aromatics, including lignin precursors. Heteronuclear single-quantum coherence (HSQC) NMR spectroscopy revealed that scopoletin in cell walls was bound to lignin, leading to a reduction in lignin content and changes in its monomeric composition and molar mass distribution. Furthermore, the enzymatic saccharification efficiency of the transgenic cell walls was more than three times higher than that of the wild-type plants, even without pretreatment. Although addressing growth inhibition remains a priority, incorporating scopoletin into lignin demonstrates significant potential for improving woody biomass utilization.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS2002

Predicting Print-thru for the Sub-scale Beryllium Mirror Demonstrator (SBMD)

This document presents a finite element method for predicting print-thru or quilting for a lightweight mirror in a low temperature environment. The mirror is represented with quadrilateral and triangular plate finite elements. The SBMD (Sub-scale Beryllium Mirror Demonstrator) is circular with a diameter of 50 cm and one flat side. The mirror structure is a thin-wall triangular cell core with a single facesheet. There is a 4 mm radius fillet between the facesheet and cell walls. It is made entirely of Beryllium. It is assumed that polishing the mirror surface creates a thin surface layer with different material properties. Finite element results are compared with measured values at cryogenic temperatures.

Craig, Larry↗