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At least 307 records · Page 17

The evolution of the Human Systems and Simulation Laboratory in nuclear power research

The events at Three Mile Island in the United States brought about fundamental changes in the ways that simulation would be used in nuclear operations. The need for research simulators was identified to scientifically study human-centered risk and make recommendations for process control system designs. This paper documents the human factors research conducted at the Human Systems and Simulation Laboratory (HSSL) since its inception in 2010 at Idaho National Laboratory. The facility’s primary purposes are to provide support to utilities for system upgrades and to validate modernized control room concepts. In the last decade, however, as nuclear industry needs have evolved, so too have the purposes of the HSSL. Thus, beyond control room modernization, human factors researchers have evaluated the security of nuclear infrastructure from cyber adversaries and evaluated human-in-the-loop simulations for joint operations with an integrated hydrogen generation plant. Lastly, our review presents research using human reliability analysis techniques with data collected from HSSL-based studies and concludes with potential future directions for the HSSL, including severe accident management and advanced control room technologies.

21 SPECIFIC NUCLEAR REACTORS AND ASSOCIATED PLANTS↗

Three-dimensional patchy lattice model: Ring formation and phase separation

We investigate the structural and thermodynamic properties of a model of particles with 2 patches of type A and 10 patches of type B. Particles are placed on the sites of a face centered cubic lattice with the patches oriented along the nearest neighbor directions. The competition between the self-assembly of chains, rings, and networks on the phase diagram is investigated by carrying out a systematic investigation of this class of models, using an extension of Wertheim's theory for associating fluids and Monte Carlo numerical simulations. We varied the ratio r ≡ ε{sub AB}/ε{sub AA} of the interaction between patches A and B, ε{sub AB}, and between A patches, ε{sub AA} (ε{sub BB} is set to 0) as well as the relative position of the A patches, i.e., the angle θ between the (lattice) directions of the A patches. We found that both r and θ (60°, 90°, or 120°) have a profound effect on the phase diagram. In the empty fluid regime (r < 1/2) the phase diagram is reentrant with a closed miscibility loop. The region around the lower critical point exhibits unusual structural and thermodynamic behavior determined by the presence of relatively short rings. The agreement between the results of theory and simulation is excellent for θ = 120° but deteriorates as θ decreases, revealing the need for new theoretical approaches to describe the structure and thermodynamics of systems dominated by small rings.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

SUMOylation of KLF4 acts as a switch in transcriptional programs that control VSMC proliferation

The regulation of vascular smooth muscle cell (VSMC) proliferation is an important issue due to its major implications for the prevention of pathological vascular conditions. The objective of this work was to assess the function of small ubiquitin-like modifier (SUMO)ylated Krϋppel-like transcription factor 4 (KLF4) in the regulation of VSMC proliferation in cultured cells and in animal models with balloon injury. We found that under basal conditions, binding of non-SUMOylated KLF4 to p300 activated p21 (p21{sup WAF1/CIP1})transcription, leading to VSMC growth arrest. PDGF-BB promoted the interaction between Ubc9 and KLF4 and the SUMOylation of KLF4, which in turn recruited transcriptional corepressors to the p21 promoter. The reduction in p21 enhanced VSMC proliferation. Additionally, the SUMOylated KLF4 did not affect the expression of KLF4, thereby forming a positive feedback loop enhancing cell proliferation. These results demonstrated that SUMOylated KLF4 plays an important role in cell proliferation by reversing the transactivation action of KLF4 on p21 induced with PDGF-BB.

60 APPLIED LIFE SCIENCES↗

RISK-INFORMED EXTERNAL HAZARDS ANALYSIS BY EEVE TOOLKIT

Introducing risk-informed safety analyses for external events can bring sensible benefits to the nuclear power industry easing the regulatory burden introduced in the aftermath of the Fukushima accident. In the framework of the US-DOE Light-Water Reactor Sustainability program, Risk-Informed Margin Characterization project (LWRS-RISMC), research activities were conducted at INL for developing and applying a toolkit for performing risk-informed external hazards analysis. The “EEVE” (External EVEnts) toolkit, a chain of INL and state-of-the-art codes, was set-up and applied for simulating earthquake-induced transients with internal flooding in a PWR. This was done using methodology set-up for evaluating new data and determining critical areas that would benefit most from advanced analysis methods. The methodology is agnostic and it can be applied to other external hazards than just seismic and flooding (e.g., high winds, intense precipitation, etc.). The first step was the deterministic calculations for evaluating the propagation of a set of earthquakes and for determining the effects on a fire suppression system in an auxiliary building. These deterministic calculations were performed using the LS-DYNA and the OPENSEE structural mechanics codes. Probability failures and event trees of a generic 3-loops Westinghouse PWR were derived from a generic SAPHIRE model. Seismically induced Loss of Offsite Power, Station Blackout, and fire suppression piping failure events were considered. SAPHIRE code allowed a pre-screening of all possible branches of the event tree and a selection of those with the largest Core Damage Frequency increase for further analysis using dynamic PRA methods. INL's EMRALD code performed dynamic PRA calculations of the identified sequences, integrating risk analysis with on-line deterministic safety analysis results (seismically-induced internal flooding calculations by NEUTRINO code and system analysis by RELAP5-3D code). RELAP5-3D Best-Estimate calculations coupled with RAVEN statistical code were used for deriving limit surfaces as function of relevant transient parameters. This data was used by EMRALD for determining the final core status (core damage/core safe). Finally, the coupling of seismic, flooding, system thermal-hydraulic, uncertainty quantification and probabilistic risk analysis for an advanced risk-informed external hazards analysis was demonstrated. Results showed the sensible decrease in the level of conservatism achieved using the developed risk-informed methodology, which allowed also improving the evaluation of the failure modes and failure probabilities of different components.

22 GENERAL STUDIES OF NUCLEAR REACTORS↗

Thermodynamic Limits of Redox-Based Thermochemical Processes (REDOTHERM)

Solar thermochemical fuel production is a potential pathway for the production of sustain liquid drop-in fuels, which can help decarbonize the aviation and maritime sectors. In an attempt to analyze the commercial viability of this technology, several studies have been conducted, including system and technoeconomic analysis (TEA) modeling. However, most studies to date simply assume a given redox reactor efficiency, which is significantly higher than demonstrated values to date. While it is widely recognized that utilizing a counter-current flow (CF) configuration could increase the redox reactor efficiency, an over-simplification in the thermodynamic modeling may lead to unphysical results which has been included in multiple publications. The fact that the solar redox reactor is the least developed component in the process chain makes it hard to identify technology gaps and evaluate pathways to deployment at scale using this approach. In this work, a thermodynamic model for a moving oxide system has been developed, in a general form that allows to analyze the system for different redox-active materials, under a wide range of operating conditions, for both parallel and countercurrent flows. The model capabilites are demonstrated, and the model's code will be shared as an open-source on GitHub in the next few months.

chemical looping↗

Quantum field theory for the chiral clock transition in one spatial dimension

We describe the quantum phase transition in the N-state chiral clock model in spatial dimension d = 1. With couplings chosen to preserve time-reversal and spatial inversion symmetries, such a model is in the universality class of recent experimental studies of the ordering of pumped Rydberg states in a one-dimensional chain of trapped ultracold alkali atoms. For such couplings and N = 3, the clock model is expected to have a direct phase transition from a gapped phase with a broken global Z N symmetry, to a gapped phase with the Z N symmetry restored. The transition has dynamical critical exponent z ≠ 1, and so cannot be described by a relativistic quantum field theory. We use a lattice duality transformation to map the transition onto that of a Bose gas in d = 1, involving the onset of a single-boson condensate in the background of a higher-dimensional N-boson condensate. Here, we present a renormalization group analysis of the strongly coupled field theory for the Bose gas transition in an expansion in 2 – d, with 4 – N chosen to be of order 2 – d. At two-loop order, we find a regime of parameters with a renormalization group fixed point which can describe a direct phase transition. We also present numerical density-matrix renormalization group studies of lattice chiral clock and Bose gas models for N = 3, finding good evidence for a direct phase transition, and obtain estimates for z and the correlation length exponent ν.

75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND↗

Long none coding RNA HOTTIP/HOXA13 act as synergistic role by decreasing cell migration and proliferation in Hirschsprung disease

Long noncoding RNAs (lncRNAs) have been confirmed to be associated with various human diseases. However, whether they are associated with Hirschsprung disease (HSCR) progression remains unclear. In this study, we designed the experiment to explore the relationship between lncRNA HOTTIP and HOXA13, and their pathogenicity to HSCR. Quantitative real-time PCR and Western blot were performed to detect the levels of lncRNA, mRNAs, and proteins in colon tissues from 79 patients with HSCR and 79 controls. Small RNA interference transfection was used to study the function experiments in human 293T and SK-N-BE cell lines. The cell viability and activities were detected by the transwell assays, CCK8 assay, and flow cytometry, respectively. LncRNA HOTTIP and HOXA13 were significantly down-regulated in HSCR compared to the controls. Meanwhile, the declined extent of their expression levels makes sense between two main phenotype of HSCR. SiRNA-mediated knock-down of HOTTIP or HOXA13 correlated with decreased levels of each other and both reduced the cell migration and proliferation without affecting cell apoptosis or cell cycle. Our study demonstrates that aberrant reduction of HOTTIP and HOXA13, which have a bidirectional regulatory loop, may play an important role in the pathogenesis of HSCR. - Highlights: • LncRNA HOTTIP and HOXA13 are both down-regulated in HSCR. • HOTTIP and HOXA13 can regulate each other in 293T and SK-N-BE(2) cell lines. • Both HOTTIP and HOXA13 can decrease cell migration and proliferation.

60 APPLIED LIFE SCIENCES↗

Solute-solvent cavity and bridge functions. I. Varying size of the solute

In this work we present the results of the extensive molecular simulations of solute-solvent cavity and bridge functions. The mixtures of Lennard-Jones solvent with Lennard-Jones solute at infinite dilution are considered for different solute-solvent size ratios—up to 4:1. The Percus-Yevick and hypernetted chain closures deviate substantially from simulation results in the investigated temperature and density ranges. We also find that the behavior of the indirect and cavity correlation functions is non-monotonous within the hard-core region, but the latter can be successfully approximated by mean-field theory if the solute-solvent interaction energy is divided into repulsive and attractive contribution, according to Weeks-Chandler-Andersen theory. Furthermore, in spite of the non-monotonous behavior of logarithm of the cavity function and the indirect correlation function, their difference, i.e., the bridge function remains constant within the hard-core region. Such behavior of the bridge and indirect correlation functions at small distances and for small values of indirect correlation function is well known from the Duh-Haymet plots, where the non-unique relationship results in loops of the bridge function vs. indirect correlation function graphs. We show that the same pathological behavior appears also when distance is small and indirect correlation function is large. We further show that the unique functional behavior of the bridge function can be established when bridge is represented as a function of the renormalized, repulsive indirect correlation function.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

A plasmid borne, functionally novel glycoside hydrolase family 30 subfamily 8 endoxylanase from solventogenic Clostridium

Glycoside hydrolase family 30 subfamily 8 (GH30-8) β-1,4-endoxylanases are known for their appendage-dependent function requiring recognition of an α-1,2-linked glucuronic acid (GlcA) common to glucuronoxylans for hydrolysis. Structural studies have indicated that the GlcA moiety of glucuronoxylans is coordinated through six hydrogen bonds and a salt bridge. These GlcA-dependent endoxylanases do not have significant activity on xylans that do not bear GlcA substitutions such as unsubstituted linear xylooligosaccharides or cereal bran arabinoxylans. In the present study, we present the structural and biochemical characteristics of xylanase 30A from Clostridium acetobutylicum (CaXyn30A) which was originally selected for study due to predicted structural differences within the GlcA coordination loops. Amino acid sequence comparisons indicated that this Gram-positive-derived GH30-8 more closely resembles Gram-negative derived forms of these endoxylanases: a hypothesis borne out in the developed crystallographic structure model of the CaXyn30A catalytic domain (CaXyn30A-CD). CaXyn30A-CD hydrolyzes xylans to linear and substituted oligoxylosides showing the greatest rate with the highly arabinofuranose (Araf)-substituted cereal arabinoxylans. CaXyn30A-CD hydrolyzes xylooligosaccharides larger than xylotriose and shows an increased relative rate of hydrolysis for xylooligosaccharides containing α-1,2-linked arabinofuranose substitutions. Biochemical analysis confirms that CaXyn30A benefits from five xylose-binding subsites which extend from the -3 subsite to the +2 subsite of the binding cleft. These studies indicate that CaXyn30A is a GlcA-independent endoxylanase that may have evolved for the preferential recognition of α-1,2-Araf substitutions on xylan chains.

59 BASIC BIOLOGICAL SCIENCES↗

Crystal Structure of the Receptor Binding Domain of the botulinum C-D Mosiac Neurotoxin Reveals Potential Roles of Lysines 1118 and 1136 in Membrane Interactions

The botulinum neurotoxins (BoNTs) produced by different strains of the bacterium Clostridium botulinum are responsible for the disease botulism and include a group of immunologically distinct serotypes (A, B, E, and F) that are considered to be the most lethal natural proteins known for humans. Two BoNT serotypes, C and D, while rarely associated with human infection, are responsible for deadly botulism outbreaks afflicting animals. Also associated with animal infections is the BoNT C-D mosaic protein (BoNT/CD), a BoNT subtype that is essentially a hybrid of the BoNT/C ({approx}two-third) and BoNT/D ({approx}one-third) serotypes. While the amino acid sequence of the heavy chain receptor binding (HCR) domain of BoNT/CD (BoNT/CD-HCR) is very similar to the corresponding amino acid sequence of BoNT/D, BoNT/CD-HCR binds synaptosome membranes better than BoNT/D-HCR. To obtain structural insights for the different membrane binding properties, the crystal structure of BoNT/CD-HCR (S867-E1280) was determined at 1.56 {angstrom} resolution and compared to previously reported structures for BoNT/D-HCR. Overall, the BoNT/CD-HCR structure is similar to the two sub-domain organization observed for other BoNT HCRs: an N-terminal jellyroll barrel motif and a C-terminal {beta}-trefoil fold. Comparison of the structure of BoNT/CD-HCR with BoNT/D-HCR indicates that K1118 has a similar structural role as the equivalent residue, E1114, in BoNT/D-HCR, while K1136 has a structurally different role than the equivalent residue, G1132, in BoNT/D-HCR. Lysine-1118 forms a salt bridge with E1247 and may enhance membrane interactions by stabilizing the putative membrane binding loop (K1240-N1248). Lysine-1136 is observed on the surface of the protein. A sulfate ion bound to K1136 may mimic a natural interaction with the negatively changed phospholipid membrane surface. Liposome-binding experiments demonstrate that BoNT/CD-HCR binds phosphatidylethanolamine liposomes more tightly than BoNT/D-HCR.

59 BASIC BIOLOGICAL SCIENCES↗

Renormalization group study of the minimal Majoronic dark radiation and dark matter model

We study the 1-loop renormalization group equation running in the simplest singlet Majoron model constructed by us earlier to accommodate the dark radiation and dark matter content in the universe. A comprehensive numerical study was performed to explore the whole model parameter space. A smaller effective number of neutrinos △N{sub eff}∼0.05, or a Majoron decoupling temperature higher than the charm quark mass, is preferred. We found that a heavy scalar dark matter, ρ, of mass 1.5–4 TeV is required by the stability of the scalar potential and an operational type-I see-saw mechanism for neutrino masses. A neutral scalar, S, of mass in the 10–100 GeV range and its mixing with the standard model Higgs as large as 0.1 is also predicted. The dominant decay modes are S into bb-bar and/or ωω. A sensitive search will come from rare Z decays via the chain Z→S+ff-bar, where f is a Standard Model fermion, followed by S into a pair of Majoron and/or b-quarks. The interesting consequences of dark matter bound state due to the sizable Sρρ-coupling are discussed as well. In particular, shower-like events with an apparent neutrino energy at M{sub ρ} could contribute to the observed effective neutrino flux in underground neutrino detectors such as IceCube.

71 CLASSICAL AND QUANTUM MECHANICS, GENERAL PHYSIC↗

Inheritance of mitochondrial DNA in serially recloned pigs by somatic cell nuclear transfer (SCNT)

Highlights: Black-Right-Pointing-Pointer We success serial SCNT through the third generation using pig fibroblasts. Black-Right-Pointing-Pointer Donor-specific mtDNA in the recloned pigs was detected. Black-Right-Pointing-Pointer SCNT affect mtDNA mounts. -- Abstract: Somatic cell nuclear transfer (SCNT) has been established for the transmission of specific nuclear DNA. However, the fate of donor mitochondrial DNA (mtDNA) remains unclear. Here, we examined the fate of donor mtDNA in recloned pigs through third generations. Fibroblasts of recloned pigs were obtained from offspring of each generation produced by fusion of cultured fibroblasts from a Minnesota miniature pig (MMP) into enucleated oocytes of a Landrace pig. The D-loop regions from the mtDNA of donor and recipient differ at nucleotide sequence positions 16050 (A{yields}T), 16062 (T{yields}C), and 16135 (G{yields}A). In order to determine the fate of donor mtDNA in recloned pigs, we analyzed the D-loop region of the donor's mtDNA by allele-specific PCR (AS-PCR) and real-time PCR. Donor mtDNA was successfully detected in all recloned offspring (F1, F2, and F3). These results indicate that heteroplasmy that originate from donor and recipient mtDNA is maintained in recloned pigs, resulting from SCNT, unlike natural reproduction.

60 APPLIED LIFE SCIENCES↗

KRAS Switch Mutants D33E and A59G Crystallize in the State 1 Conformation

KRAS switch loop movements play a crucial role in regulating RAS signaling, and alteration of these sensitive dynamics is a principal mechanism through which disease-associated RAS mutations lead to aberrant RAS activation. Prior studies suggest that despite a high degree of sequence similarity, the switches in KRAS are more dynamic than those in HRAS. We determined X-ray crystal structures of the rare tumorigenic KRAS mutants KRAS D33E , in switch 1 (SW1), and KRAS A59G , in switch 2 (SW2), bound to GDP and found these adopt nearly identical, open SW1 conformations as well as altered SW2 conformations. KRAS A59G bound to a GTP analogue crystallizes in the same conformation. This open conformation is consistent with the inactive “state 1” previously observed for HRAS bound to GTP. For KRAS A59G , switch rearrangements may be regulated by increased flexibility in the 57 DXXGQ 61 motif at codon 59. However, loss of interactions between side chains at codons 33 and 35 in the SW1 33 DPT 35 motif drives changes for KRAS D33E . The 33 DPT 35 motif is conserved for multiple members of the RAS subfamily but is not found in RAB, RHO, ARF, or Gα families, suggesting that dynamics mediated by this motif may be important for determining the selectivity of RAS–effector interactions. Biochemically, the consequence of altered switch dynamics is the same, showing impaired interaction with the guanine exchange factor SOS and loss of GAP-dependent GTPase activity. However, interactions with the RBD of RAF are preserved. Furthermore, these observations add to a body of evidence suggesting that HRAS and KRAS show meaningful differences in functionality stemming from differential protein dynamics independent of the hypervariable region.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Plant-specific features of respiratory supercomplex I + III2 from Vigna radiata

Abstract The last steps of cellular respiration—an essential metabolic process in plants—are carried out by mitochondrial oxidative phosphorylation. This process involves a chain of multi-subunit membrane protein complexes (complexes I–V) that form higher-order assemblies called supercomplexes. Although supercomplexes are the most physiologically relevant form of the oxidative phosphorylation complexes, their functions and structures remain mostly unknown. Here we present the cryogenic electron microscopy structure of the supercomplex I + III 2 from Vigna radiata (mung bean). The structure contains the full subunit complement of complex I, including a newly assigned, plant-specific subunit. It also shows differences in the mitochondrial processing peptidase domain of complex III 2 relative to a previously determined supercomplex with complex IV. The supercomplex interface, while reminiscent of that in other organisms, is plant specific, with a major interface involving complex III 2 ’s mitochondrial processing peptidase domain and no participation of complex I’s bridge domain. The complex I structure suggests that the bridge domain sets the angle between the enzyme’s two arms, limiting large-scale conformational changes. Moreover, complex I’s catalytic loops and its response in active-to-deactive assays suggest that, in V. radiata , the resting complex adopts a non-canonical state and can sample deactive- or open-like conformations even in the presence of substrate. This study widens our understanding of the possible conformations and behaviour of complex I and supercomplex I + III 2 . Further studies of complex I and its supercomplexes in diverse organisms are needed to determine the universal and clade-specific mechanisms of respiration.

59 BASIC BIOLOGICAL SCIENCES↗

Detection of PIWI and piRNAs in the mitochondria of mammalian cancer cells

Highlights: • piRNA sequences were mapped to human mitochondrial (mt) genome. • We inspected small RNA-Seq datasets from somatic cell mt subcellular fractions. • Piwi and piRNA transcripts are present in mammalian somatic cancer cell mt fractions. - Abstract: Piwi-interacting RNAs (piRNAs) are 26–31 nt small noncoding RNAs that are processed from their longer precursor transcripts by Piwi proteins. Localization of Piwi and piRNA has been reported mostly in nucleus and cytoplasm of higher eukaryotes germ-line cells, where it is believed that known piRNA sequences are located in repeat regions of nuclear genome in germ-line cells. However, localization of PIWI and piRNA in mammalian somatic cell mitochondria yet remains largely unknown. We identified 29 piRNA sequence alignments from various regions of the human mitochondrial genome. Twelve out 29 piRNA sequences matched stem-loop fragment sequences of seven distinct tRNAs. We observed their actual expression in mitochondria subcellular fractions by inspecting mitochondrial-specific small RNA-Seq datasets. Of interest, the majority of the 29 piRNAs overlapped with multiple longer transcripts (expressed sequence tags) that are unique to the human mitochondrial genome. The presence of mature piRNAs in mitochondria was detected by qRT-PCR of mitochondrial subcellular RNAs. Further validation showed detection of Piwi by colocalization using anti-Piwil1 and mitochondria organelle-specific protein antibodies.

60 APPLIED LIFE SCIENCES↗

Structural and Thermodynamic Comparison of the Catalytic Domain of AMSH and AMSH-LP: Nearly Identical Fold but Different Stability

AMSH plays a critical role in the ESCRT (endosomal sorting complexes required for transport) machinery, which facilitates the down-regulation and degradation of cell-surface receptors. It displays a high level of specificity toward cleavage of Lys63-linked polyubiquitin chains, the structural basis of which has been understood recently through the crystal structure of a highly related, but ESCRT-independent, protein AMSH-LP (AMSH-like protein). We have determined the X-ray structure of two constructs representing the catalytic domain of AMSH: AMSH244, the JAMM (JAB1/MPN/MOV34)-domain-containing polypeptide segment from residues 244 to 424, and AMSH219{sup E280A}, an active-site mutant, Glu280 to Ala, of the segment from 219 to 424. In addition to confirming the expected zinc coordination in the protein, the structures reveal that the catalytic domains of AMSH and AMSH-LP are nearly identical; however, guanidine-hydrochloride-induced unfolding studies show that the catalytic domain of AMSH is thermodynamically less stable than that of AMSH-LP, indicating that the former is perhaps structurally more plastic. Much to our surprise, in the AMSH219{sup E280A} structure, the catalytic zinc was still held in place, by the compensatory effect of an aspartate from a nearby loop moving into a position where it could coordinate with the zinc, once again suggesting the plasticity of AMSH. Additionally, a model of AMSH244 bound to Lys63-linked diubiquitin reveals a type of interface for the distal ubiquitin significantly different from that seen in AMSH-LP. Altogether, we believe that our data provide important insight into the structural difference between the two proteins that may translate into the difference in their biological function.

Davies, Christopher W.↗

Perturbation of the Monomer-Monomer Interfaces of the Benzoylformate Decarboxylase Tetramer

The X-ray structure of benzoylformate decarboxylase (BFDC) from Pseudomonas putida ATCC 12633 shows it to be a tetramer. This was believed to be typical of all thiamin diphosphate-dependent decarboxylases until recently when the structure of KdcA, a branched-chain 2-keto acid decarboxylase from Lactococcus lactis, showed it to be a homodimer. This lent credence to earlier unfolding experiments on pyruvate decarboxylase from Saccharomyces cerevisiae that indicated that it might be active as a dimer. To investigate this possibility in BFDC, we sought to shift the equilibrium toward dimer formation. Point mutations were made in the noncatalytic monomer–monomer interfaces, but these had a minimal effect on both tetramer formation and catalytic activity. Subsequently, the R141E/Y288A/A306F variant was shown by analytical ultracentrifugation to be partially dimeric. It was also found to be catalytically inactive. Further experiments revealed that just two mutations, R141E and A306F, were sufficient to markedly alter the dimer–tetramer equilibrium and to provide an ~450-fold decrease in k cat . Equilibrium denaturation studies suggested that the residual activity was possibly due to the presence of residual tetramer. The structures of the R141E and A306F variants, determined to <1.5 Å resolution, hinted that disruption of the monomer interfaces will be accompanied by movement of a loop containing Leu109 and Leu110. As these residues contribute to the hydrophobicity of the active site and the correct positioning of the substrate, it seems that tetramer formation may well be critical to the catalytic activity of BFDC.

Andrews, Forest H.↗

2024 Incubator Final Report: Active Hybrid Mooring

This Incubator project has defined and developed a representation of an active hybrid mooring (AHM) system in Simulink to assess whether the speed and accuracy of AHM is sufficient to warrant further investigation in a larger project. We first built a representative mooring system for the VolturnUS-S 15MW floating offshore wind turbine (FOWT) in 1,000-meter water depth to provide baseline loads and displacements needed in the AHM system, with the results showing off-the-shelf actuators would be practical. We then defined model configurations for the numerical and physical substructures, using standalone MoorDyn v2 with a novel input strategy and chained hydraulic actuators represented in Simulink Simscape, respectively. The remainder of the project has focused on verifying the existing models for speed and accuracy against OpenFAST simulations, increasing their complexity to enable six degree of freedom (6DOF) operation, and connecting the numerical and physical substructures into a single holistic model. The verification phase of the project has shown promising results, though further refinement is needed to resolve accuracy discrepancies and for robust performance across a wider range of metocean conditions. The numerical model we developed is computationally stable and operates faster than real time using a time step of 0.01 seconds or shorter, indicating the model is likely fast enough for our AHM design to operate successfully. Additionally, the Simscape model of a hydraulic actuator operates with less than 0.3% relative error when actuating mooring loads from an OpenFAST simulation of the VolturnUS-S 1,000-meter mooring system. The aforementined refinements will be performed in the next year as a Continuing Incubator, with key remaining tasks include programming the predictor-corrector loop to minimize movement error, testing against more complex nonlinear wave cases, and expanding to enable loads from connected shared mooring lines.

17 WIND ENERGY↗