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Results for “substrate selection”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 73 records · Page 4
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Data Science-Driven Analysis of Substrate-Permissive Diketopiperazine Reverse Prenyltransferase NotF: Applications in Protein Engineering and Cascade Biocatalytic Synthesis of (-)-Eurotiumin A

Prenyltransfer is an early-stage carbon-hydrogen bond (C-H) functionalization prevalent in the biosynthesis of a diverse array of biologically active bacterial, fungal, plant, and metazoan diketopiperazine (DKP) alkaloids. Toward the development of a unified strategy for biocatalytic construction of prenylated DKP indole alkaloids, we sought to identify and characterize a substrate-permissive C2 reverse prenyltransferase (PT). As the first tailoring event within the biosynthesis of cytotoxic notoamide metabolites, PT NotF catalyzes C2 reverse prenyltransfer of brevianamide F. Solving a crystal structure of NotF (in complex with native substrate and prenyl donor mimic dimethylallyl S-thiolodiphosphate (DMSPP)) revealed a large, solvent-exposed active site, intimating NotF may possess a significantly broad substrate scope. To assess the substrate selectivity of NotF, we synthesized a panel of 30 sterically and electronically differentiated tryptophanyl DKPs, the majority of which were selectively prenylated by NotF in synthetically useful conversions (2 to > 99%). Quantitative representation of this substrate library and development of a descriptive statistical model provided insight into the molecular origins of NotF's substrate promiscuity. This approach enabled the identification of key substrate descriptors (electrophilicity, size, and flexibility) that govern the rate of NotF-catalyzed prenyltransfer, and the development of an "induced fit docking (IFD)-guided" engineering strategy for improved turnover of our largest substrates. We further demonstrated the utility of NotF in tandem with oxidative cyclization using flavin monooxygenase, BvnB. This one-pot, in vitro biocatalytic cascade enabled the first chemoenzymatic synthesis of the marine fungal natural product, (-)-eurotiumin A, in three steps and 60% overall yield.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2019

Structural and dynamical rationale for fatty acid unsaturation in Escherichia coli

Fatty acid biosynthesis in α- and γ-proteobacteria requires two functionally distinct dehydratases, FabA and FabZ. In this work, mechanistic cross-linking facilitates the structural characterization of a stable hexameric complex of six Escherichia coli FabZ dehydratase subunits with six AcpP acyl carrier proteins. The crystal structure sheds light on the divergent substrate selectivity of FabA and FabZ by revealing distinct architectures of the binding pocket. Molecular dynamics simulations demonstrate differential biasing of substrate orientations and conformations within the active sites of FabA and FabZ such that FabZ is preorganized to catalyze only dehydration, while FabA is primed for both dehydration and isomerization.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Catalytic polymer processing

A method of upcycling polymers to useful hydrocarbon materials. A catalyst with nanoparticles on a substrate selectively docks and cleaves longer hydrocarbon chains over shorter hydrocarbon chains. The catalyst includes metal nanoparticles in an order array on a substrate.

Delferro, Massimiliano↗
DOE OSTI2024

Principles of paralog-specific targeted protein degradation engaging the C-degron E3 KLHDC2

Abstract PROTAC® (proteolysis-targeting chimera) molecules induce proximity between an E3 ligase and protein-of-interest (POI) to target the POI for ubiquitin-mediated degradation. Cooperative E3-PROTAC-POI complexes have potential to achieve neo-substrate selectivity beyond that established by POI binding to the ligand alone. Here, we extend the collection of ubiquitin ligases employable for cooperative ternary complex formation to include the C-degron E3 KLHDC2. Ligands were identified that engage the C-degron binding site in KLHDC2, subjected to structure-based improvement, and linked to JQ1 for BET-family neo-substrate recruitment. Consideration of the exit vector emanating from the ligand engaged in KLHDC2’s U-shaped degron-binding pocket enabled generation of SJ46421, which drives formation of a remarkably cooperative, paralog-selective ternary complex with BRD3 BD2 . Meanwhile, screening pro-drug variants enabled surmounting cell permeability limitations imposed by acidic moieties resembling the KLHDC2-binding C-degron. Selectivity for BRD3 compared to other BET-family members is further manifested in ubiquitylation in vitro, and prodrug version SJ46420-mediated degradation in cells. Selectivity is also achieved for the ubiquitin ligase, overcoming E3 auto-inhibition to engage KLHDC2, but not the related KLHDC1, KLHDC3, or KLHDC10 E3s. In sum, our study establishes neo-substrate-specific targeted protein degradation via KLHDC2, and provides a framework for developing selective PROTAC protein degraders employing C-degron E3 ligases.

Science & Technology - Other Topics↗
NASA NTRS1993

High-Tc thermal bridges for space-borne cryogenic infrared detectors

The potential for using high-temperature superconductive elements, screen-printed onto ceramic substrates, as thermal bridges to replace the currently employed manganin wires is studied at NASA-LaRC. Substrate selection is considered to be the most critical parameter in device production. Due to the glass-like thermal behavior of yttria-stabilized-zirconia (YSZ) and fused silica substrates, these materials are found to reduce the heat load significantly. The estimated thermal savings for superconductive leads printed onto YSZ or fused silica substrates range from 6 to 14 percent.

Wise, S. A.↗
DOE OSTI2019

Insight into subtilisin E-S7 cleavage pattern based on crystal structure and hydrolysates peptide analysis

The X-ray crystallographic structure of the mature form of subtilisin E-S7 (SES7) at 1.90 Å resolution is reported in this paper. Structural comparisons between the previously reported propeptide-subtilisin E complex (1SCJ) and our mature form subtilisin E-S7 (6O44) provide insight into active site adjustments involved in catalysis and specificity. To further investigate the protease substrate selectivity mechanism, we used SES7 to hydrolyze skim milk and analyzed the hydrolysates by LC-MS for peptide identification. The cleavage pattern suggests a high preference for proline at substrate P2 position. The results based on the peptide analysis are consistent with our structural observations, which is instrumental in future protein engineering by rational design. Furthermore, the ACE-inhibitor and NLN-inhibitor activity of the hydrolysates were determined to assess the utility of SES7 for further industrial applications; IC50-ACE = 67 ± 0.92 μg/mL and IC50-NLN = 263 ± 13 μg/mL.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Arsinothricin, an arsenic-containing non-proteinogenic amino acid analog of glutamate, is a broad-spectrum antibiotic

Abstract The emergence and spread of antimicrobial resistance highlights the urgent need for new antibiotics. Organoarsenicals have been used as antimicrobials since Paul Ehrlich’s salvarsan. Recently a soil bacterium was shown to produce the organoarsenical arsinothricin. We demonstrate that arsinothricin, a non-proteinogenic analog of glutamate that inhibits glutamine synthetase, is an effective broad-spectrum antibiotic against both Gram-positive and Gram-negative bacteria, suggesting that bacteria have evolved the ability to utilize the pervasive environmental toxic metalloid arsenic to produce a potent antimicrobial. With every new antibiotic, resistance inevitably arises. The arsN1 gene, widely distributed in bacterial arsenic resistance ( ars ) operons, selectively confers resistance to arsinothricin by acetylation of the α-amino group. Crystal structures of ArsN1 N -acetyltransferase, with or without arsinothricin, shed light on the mechanism of its substrate selectivity. These findings have the potential for development of a new class of organoarsenical antimicrobials and ArsN1 inhibitors.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2018

AM metal substrates for inkjet-printing of smart devices

Highlights: • An experimental study to characterize 3D-printed metal substrates and their surfaces for inkjet-printing is presented. • Multilayer structures composed of an insulating layer and a superposed conductive layer, inkjet-printed onto various metallic substrates. • Characterization based on microscopic and Focused Ion Beam (FIB) images, profilometer and contact angle measurements of the substrates. • Microscopic images and four-point-probe resistance- and profilometer measurements to gain insight into the quality of the multilayer print. - Abstract: A study to characterize surface and structural properties of 3D-printed metal substrates for multilayer inkjet-printing is presented. Such substrates can enable and alleviate the fabrication of smart and functional packages, implants or spare parts. For the characterization, a multilayer structure composed of an insulating layer and a superposed conductive layer is inkjet-printed onto the various 3D-printed metallic substrates. The printing is done using equivalent settings for all substrates. The characterization is based on microscopic and Focused Ion Beam (FIB) images, as well as profilometer and contact angle measurements of the substrates. Furthermore, microscopic images and four-point-probe resistance- and profilometer measurements are carried out, to gain insight into the quality of the multilayer print. We show the diversity in surface and structural properties of 3D-printed substrates, also for those fabricated using the same process. Also, the 3D-printed samples exhibit different wettability properties than metals processed using standard processes. Finally, we can also demonstrate the suitability of selected substrates for inkjet-printing.

36 MATERIALS SCIENCE↗
DOE OSTI2025

Unique Structural Features Relate to Evolutionary Adaptation of Cytochrome P450 in the Abyssal Zone

Cytochromes P450 (CYPs) form one of the largest enzyme superfamilies, with similar structural folds yet biological functions varying from synthesis of physiologically essential compounds to metabolism of myriad xenobiotics. Sterol 14α-demethylases (CYP51s) represent a very special P450 family, regarded as a possible evolutionary progenitor for all currently existing P450s. In metazoans CYP51 is critical for the biosynthesis of sterols including cholesterol. Here we determined the crystal structures of ligand-free CYP51s from the abyssal fish Coryphaenoides armatus and human-. Comparative sequence–structure–function analysis revealed specific structural elements that imply elevated conformational flexibility, uncovering a molecular basis for faster catalytic rates, lower substrate selectivity, and intrinsic resistance to inhibition. In addition, the C. armatus structure displayed a large-scale repositioning of structural segments that, in vivo, are immersed in the endoplasmic reticulum membrane and border the substrate entrance (the FG arm, >20 Å, and the β4 hairpin, >15 Å). The structural distinction of C. armatus CYP51, which is the first structurally characterized deep sea P450, suggests stronger involvement of the membrane environment in regulation of the enzyme function. We interpret this as a co-adaptation of the membrane protein structure with membrane lipid composition during evolutionary incursion to life in the deep sea.

Biochemistry & Molecular Biology↗
DOE OSTI2017

Critical Role of Surface Energy in Guiding Crystallization of Solution-Coated Conjugated Polymer Thin Films

It is well-known that substrate surface properties have a profound impact on morphology of thin films solution coated atop and the resulting solid-state properties. However, design rules for guiding the substrate selection have not yet been established. Such design rules are particularly important for solution coated semiconducting polymers, as the substratedirected thin film morphology can impact charge transport properties by orders of magnitude. We hypothesize that substrate surface energies dictate the thin film morphology by modulating the free energy barrier to heterogeneous nucleation. To test this hypothesis, we systematically vary the substrate surface energy via surface functionalization techniques. We perform in-depth morphology and device characterizations to establish the relationship between substrate surface energy, thin film morphology and charge transport properties, employing a donor-accepter (D-A) conjugated polymer. Here, we find that decreasing the substrate surface energy progressively increases thin film crystallinity, degree of molecular ordering and extent of domain alignment. Notably, the enhanced morphology on the lowest surface energy substrate lead to a 10-fold increase in the charge carrier mobility. We further develop a free energy model relating the substrate surface energy to the penalty of heterogeneous nucleation from solution in the thin film geometry. The model correctly predicts the experimental trend, thereby validating our hypothesis. This work is a significant step towards establishing design rules and understanding the critical role of substrates in determining morphology of solution coated thin films.

36 MATERIALS SCIENCE↗
DOE OSTI2020

Bifunctional Heterometallic Metal-Organic Frameworks for Solvent-Free Heterogeneous Cascade Catalysis

A family of heterometallic metal-organic frameworks (MOFs) (CPM200s) harmoniously coexisting as Lewis acids and base (azo) sites were prepared. Seven CPM200s were employed as multifunctional heterogeneous cascade catalysts for the one-pot deacetalization-Knoevenagel reaction in a solvent-free system. Benefiting from the cooperation between Lewis acids from the open metal sites and base sites from the ligands, the CPM200s showed high activity and selectivity for the tandem reaction. The heterometallic 3D porous framework reported here not only offers a combination of two opposite active sites in the same framework of materials but also increases mass transfer of the substrate, thus maximizing the efficiency and substrate selectivity of the bifunctional catalysts. The CPM200s showed the highest turnover frequency (TOF), outperforming that of the reported MOFs in tandem with the deacetalization-Knoevenagel reaction. A strong correlation between the TOF and charge-to-radius ratio (z/r) of metal ions in the CPM200s was observed for the first time. The bifunctional CPM200s catalysts can be reused five times without significant loss of activity.

36 MATERIALS SCIENCE↗
DOE OSTI2023

Structural basis of promiscuous substrate transport by Organic Cation Transporter 1

Organic Cation Transporter 1 (OCT1) plays a crucial role in hepatic metabolism by mediating the uptake of a range of metabolites and drugs. Genetic variations can alter the efficacy and safety of compounds transported by OCT1, such as those used for cardiovascular, oncological, and psychological indications. Despite its importance in drug pharmacokinetics, the substrate selectivity and underlying structural mechanisms of OCT1 remain poorly understood. Here, we present cryo-EM structures of full-length human OCT1 in the inward-open conformation, both ligand-free and drug-bound, indicating the basis for its broad substrate recognition. Comparison of our structures with those of outward-open OCTs provides molecular insight into the alternating access mechanism of OCTs. We observe that hydrophobic gates stabilize the inward-facing conformation, whereas charge neutralization in the binding pocket facilitates the release of cationic substrates. These findings provide a framework for understanding the structural basis of the promiscuity of drug binding and substrate translocation in OCT1.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2020

The structural basis of promiscuity in small multidrug resistance transporters

By providing broad resistance to environmental biocides, transporters from the small multidrug resistance (SMR) family drive the spread of multidrug resistance cassettes among bacterial populations. A fundamental understanding of substrate selectivity by SMR transporters is needed to identify the types of selective pressures that contribute to this process. Using solid-supported membrane electrophysiology, we find that promiscuous transport of hydrophobic substituted cations is a general feature of SMR transporters. To understand the molecular basis for promiscuity, we solved X-ray crystal structures of a SMR transporter Gdx-Clo in complex with substrates to a maximum resolution of 2.3 Å. These structures confirm the family’s extremely rare dual topology architecture and reveal a cleft between two helices that provides accommodation in the membrane for the hydrophobic substituents of transported drug-like cations.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

A novel 5′-hydroxyl dinucleotide hydrolase activity for the DXO/Rai1 family of enzymes

Abstract Modifications at the 5′-end of RNAs play a pivotal role in determining their fate. In eukaryotes, the DXO/Rai1 family of enzymes removes numerous 5′-end RNA modifications, thereby regulating RNA turnover. Mouse DXO catalyzes the elimination of incomplete 5′-end caps (including pyrophosphate) and the non-canonical NAD+ cap on mRNAs, and possesses distributive 5′-3′ exoribonuclease activity toward 5′-monophosphate (5′-PO4) RNA. Here, we demonstrate that DXO also catalyzes the hydrolysis of RNAs bearing a 5′-hydroxyl group (5′-OH RNA). The crystal structure of DXO in complex with a 5′-OH RNA substrate mimic at 2.0 Å resolution provides elegant insight into the molecular mechanism of this activity. More importantly, the structure predicts that DXO first removes a dinucleotide from 5′-OH RNA. Our nuclease assays confirm this prediction and demonstrate that this 5′-hydroxyl dinucleotide hydrolase (HDH) activity for DXO is higher than the subsequent 5′-3′ exoribonuclease activity for selected substrates. Fission yeast Rai1 also has HDH activity although it does not have 5′-3′ exonuclease activity, and the Rat1-Rai1 complex can completely degrade 5′-OH RNA. An Arabidopsis DXO1 variant is active toward 5′-OH RNA but prefers 5′-PO4 RNA. Collectively, these studies demonstrate the diverse activities of DXO/Rai1 and expands the collection of RNA substrates that can undergo 5′-3′ mediated decay.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Impact of Fermentation-Derived Substrates on Hydrogen Production in Zero-Gap Microbial Electrolysis Cells

Zero-gap microbial electrolysis cells (MECs) represent a promising platform for hydrogen production from liquid waste streams due to reduced interelectrode spacing that lowers internal resistance and enhances mass transport. However, the performance and stability of zero-gap MECs treating chemically complex feedstocks remain insufficiently characterized. Here, we operated zero-gap MECs with real, unamended, corn stover dark fermentation effluent containing a wide range of organic substrates. The MECs fed fermentation effluent achieved a maximum current density of 24 A/m2 (15+-6 A/m2 over the cycle) and a hydrogen production rate of 75 L/L-d (42+-19 L/L-d over the cycle). The substrates were consumed at different rates, indicating substrate-selective utilization by the anodic microbial community. Acetate supported high and stable current generation, whereas ethanol, formate, lactate, and amino acids induced varying degrees of inhibition depending on their concentration. Residual sugars caused pronounced current fluctuations, consistent with ongoing fermentation and local pH changes. A diverse microbial community was crucial for efficiently utilizing complex organics and maximizing electrochemical performance. These results demonstrate how and to what extent substrate composition regulates zero-gap MEC performance and that microbial community and operational conditions can be leveraged to enhance performance. These novel findings provide practical guidance for achieving robust hydrogen recovery from chemically heterogeneous real liquid waste streams.

08 HYDROGEN↗
DOE OSTI2018

Structural and biochemical characterization of the yeast HD domain containing protein YGK1 reveals a metal-dependent nucleoside 5-monophosphatase

Highlights: • The yeast HD domain proteins YGK1 and YB92 as 5-nucleotidases were determined. • Extensive conserved residues in the cleft were identified essential for YGK1 activity. • YGK1 dimeric core represents a characteristic of the HD family of 5-nucleotidase. • Flexible loops might confer selectivity for nucleotide. HD-domain is a conserved domain, with the signature of histidine and aspartic (HD) residues doublets. HD-domain proteins may possess nucleotidase and phosphodiesterase activities, and they play important roles in signaling and nucleotide metabolism. In yeast, HD-domain proteins with nucleotidase activity remained unexplored. Here, we biochemically and structurally characterized two HD domain proteins YGK1 (YGL101W) and YB92 (YBR242W) from Saccharomyces cerevisiae as nucleoside 5-monophosphatases, with substrate preference for deoxyribonucleoside 5-monophosphatase over ribonucleoside 5-monophosphatase. By determining the crystal structure of YGK1, we unveiled that YGK1 structure resembled as the crystal structure of YfbR from E. coli. Size-exclusion chromatography and crosslinking studies suggested that YGK1 and YB92 existed in the form of a dimer, respectively, which were consistent with structural observation of YGK1. Site-directed mutagenesis demonstrated that more extensive conserved residues near the divalent metal coordinating active site were essential for YGK1 activity than previous suggested. The metal coordinating His89 and Asp90, and the neighboring conserved Glu93, Glu114 and Glu145 were individually critical for catalysis. In addition, alignments suggested that three flexible loops with hydrophobic residues might be implicated in substrate selectivity to nucleoside moiety. Together, our comparative structural and mutational studies suggested that YGK1 and YB92 functioned as 5-nucleotidases in S. cerevisiae.

60 APPLIED LIFE SCIENCES↗
NASA NTRS1990

Method of fabricating germanium and gallium arsenide devices

A method of semiconductor diode fabrication is disclosed which relies on the epitaxial growth of a precisely doped thickness layer of gallium arsenide or germanium on a semi-insulating or intrinsic substrate, respectively, of gallium arsenide or germanium by either molecular beam epitaxy (MBE) or by metal-organic chemical vapor deposition (MOCVD). The method involves: depositing a layer of doped or undoped silicon dioxide on a germanium or gallium arsenide wafer or substrate, selectively removing the silicon dioxide layer to define one or more surface regions for a device to be fabricated thereon, growing a matched epitaxial layer of doped germanium or gallium arsenide of an appropriate thickness using MBE or MOCVD techniques on both the silicon dioxide layer and the defined one or more regions; and etching the silicon dioxide and the epitaxial material on top of the silicon dioxide to leave a matched epitaxial layer of germanium or gallium arsenide on the germanium or gallium arsenide substrate, respectively, and upon which a field effect device can thereafter be formed.

Jhabvala, Murzban↗
DOE OSTI2025

Isolation of a Terminal Cobalt Nitride in a Metal–Organic Framework

Transition metal nitrides are reactive intermediates in biological and industrial processes. Chemists have synthesized molecular model complexes of such reactive species to understand their function and electronic requirements for new applications. However, molecular chemistry can suffer from intra- and intermolecular decomposition pathways, which preclude further discovery of unknown reactive species. Metal-organic frameworks offer an opportunity for creating long-lived forms of such species with the vacuum of the pore suppressing degradation while simultaneously enabling substrate access for controlled reactivity studies. Here, in this study, we report the characterization of an elusive terminal cobalt nitride species generated through photolysis or thermolysis of a site-isolated cobalt azide within the evacuated metal-organic framework CoN 3 -MFU-4l. The first crystal structure of such a species is presented, with vibrational, X-ray absorption, and electron paramagnetic resonance spectroscopies providing further direct evidence for its formation while elucidating its electronic structure. The system additionally enables subsequent reactivity studies with selected substrates, revealing a unique ambiphilic behavior for a metal nitride species.

Börgel, Jonas [University of California, Berkeley,↗