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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 19 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2018

A transient post-translational modification of active site cysteine alters binding properties of the parkinsonism protein DJ-1

Highlights: • A fraction of bacterially expressed polyhistidine-tagged human DJ-1 tightly binds to Ni-nitrilotriacetate (Ni-NTA) column. • The color of Ni-NTA column changes from light cyan to blue violet upon tight binding to DJ-1. • An active site cysteine is post-translationally modified in the fraction of DJ-1 tightly bound to Ni-NTA. • The modificationconferring tightbinding to Ni-NTA is unstable. • Crystal structure shows the unstable modification converts to S-carboxymethylcysteine upon elution from Ni-NTA. Mutations in the human protein DJ-1 cause early onset of Parkinson's disease. A reactive cysteine residue (Cys{sup 106}) of DJ-1 is crucial for its protective function, although the underlying mechanisms are unclear. Here we show that a fraction of bacterially expressed polyhistidine-tagged human DJ-1 could not be eluted from a Ni-nitrilotriacetate (Ni-NTA) column with 150 mM imidazole. This unusually tight binding was accompanied by the appearance of blue violet color on the Ni-NTA column. We demonstrate by X-ray crystallography that Cys{sup 106} is carboxymethylated in a fraction of DJ-1 tightly bound to Ni-NTA and that the replacement of Cys{sup 106} by serine abrogates the tight binding and the appearance of blue violet color. However, carboxymethylation of purified DJ-1 is insufficient to confer the tight binding to Ni-NTA. Moreover, when eluted protein was re-applied to the Ni-NTA column, no tight binding was observed, indicating that the formation of high affinity complex with Ni-NTA depends on a transient modification of Cys{sup 106} that transforms into a Cys{sup 106}-carboxymethyl adduct upon elution from Ni-NTA. We conclude that an unknown metabolite reacts with Cys{sup 106} of DJ-1 to result in a transient post-translational modification. This modification is distinct from simple oxidation to sulfinic or sulfenic acids and confers altered binding properties to DJ-1 suggesting that it could serve as a signal for sensing oxidant stress.

60 APPLIED LIFE SCIENCES↗
NASA NTRS2016

Reconstruction of Cysteine Biosynthesis Using Engineered Cysteine-Free and Methionine-Free Enzymes

Ten of the proteinogenic amino acids can be generated abiotically while the remaining thirteen require biology for their synthesis. Paradoxically, the biosynthesis pathways observed in nature require enzymes that are made with the amino acids they produce. For example, Escherichia coli produces cysteine from serine via two enzymes that contain cysteine. Here, we substituted alternate amino acids for cysteine and also methionine, which is biosynthesized from cysteine, in serine acetyl transferase (CysE) and O-acetylserine sulfhydrylase (CysM). CysE function was rescued by cysteine-and-methionine-free enzymes and CysM function was rescued by cysteine-free enzymes. Structural modeling suggests that methionine stabilizes CysM and is present in the active site of CysM. Cysteine is not conserved among CysE and CysM protein orthologs, suggesting that cysteine is not functionally important for its own synthesis. Engineering biosynthetic enzymes that lack the amino acids being synthesized provides insights into the evolution of amino acid biosynthesis and pathways for bioengineering.

proteinogenic amino acids↗
DOE OSTI2019

Effect of Cysteine Substitutions on the Structural and Magnetic Properties of Fe{sub 3}O{sub 4}–Cysteine/RGO and Fe{sub 3}O{sub 4}/RGO–Cysteine Nanocomposites

We synthesized by electrostatic self-assembly route in basic solution Fe{sub 3}O{sub 4}–cysteine/RGO and Fe{sub 3}O{sub 4}/RGO–cysteine nanocomposites. In this method, electrostatic interaction was created via negatively charged surface of the reduced graphene oxide and reduced graphene oxide–cysteine sheets and positively charged surface of the Fe{sub 3}O{sub 4} and Fe{sub 3}O{sub 4}–Cys nanoparticles in aqueous solution. The structural and magnetic properties of the prepared samples were analyzed by x-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, transmission electron microscopy (TEM), and vibrating sample magnetometer (VSM), respectively. The dependences of magnetization as a function of applied field using a vibrating sample magnetometer exhibit S-like curves, indicating a magnetic hysteresis behavior for all samples. It clearly showed that, the nanocomposites showed a reduction of saturation magnetization to the Fe{sub 3}O{sub 4} nanoparticles.

77 NANOSCIENCE AND NANOTECHNOLOGY↗
DOE OSTI2020

Distonic radical anion species in cysteine oxidation processes

Oxidation of cysteine residues constitutes an important regulatory mechanism in the function of biological systems. Much of this behavior is controlled by the specific chemical properties of the thiol side-chain group, where reac-tions with reactive oxygen species take place. Herein, we in-vestigated the entire cysteine oxidation cycle Cys-SH ? Cys-SOnH (n = 1, 2, 3) using cryogenic negative ion photoelectron spectroscopy and quantum-chemical calculations. The conven-tional view on the first reversible oxidation step (n = 1) is as-sociated with sulfinate species. Yet our results indicate that an alternative option exists in the form of novel distonic radical anion •OS-CH2CH(NH2)-COO- with an unpaired electron on the thiol group and excess negative charge on the carboxylate. Higher order oxidation states (n = 2, 3) are thought to be asso-ciated with irreversible oxidative damage, and our results show that excess negative charge in those cases migrates to -SOn- group. Furthermore, these species are stable towards 1e oxidation, as opposed to the n = 1 case that undergoes intra-molecular proton transfer. The molecular level insights report-ed in this work provide direct spectroscopic evidences towards unique chemical versatility of Cys-sulfenic acid (Cys-SOH) in post-translational modifications of protein systems. This research was supported by the U.S. Department of Energy (DOE), Office of Science, Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences and Biosciences (X.-B. W. and M. V.) and was performed at the EMSL, a national scientific user facility sponsored by DOE's Office of Biological and Environmental Research and located at the Pacific Northwest National Laboratory. Z. B. Q. acknowledges additional support from the National Natural Science Foundation of China (Grant No. 21873003 and 21503003) and the Special Program for Applied Research on Super Computation of the NSFC-Guangdong Joint Fund (the second phase) (Grant No. U1501501) and Super Computation of Shenzhen.

Qin, Zhengbo↗
DOE OSTI2018

Participation of cysteine 30 residue in the folding process of ovalbumin evaluated in a refolding experiment using cysteine mutants

To understand the role of cysteine (SH) residues in the folding of hen ovalbumin (OVA), SH-mutated OVAs, in which each SH residue was replaced by alanine (C11A, C30A, C367A, and C382A), were prepared. SDS-PAGE analysis under non-reducing conditions showed that the C11A and C30A mutants produced a disulfide (SS) isomer in addition to a protein with a native SS bond (Cys73–Cys120). The susceptibility to elastase digestion suggested that the Cys73 residue in the SS isomer participates as a counterpart of the SS bond. Upon refolding of the SH-mutated OVAs under the denatured and SS-reduced states, only C30A failed to refold into an intact form. This indicated that the Cys30 residue plays an important role in correct refolding. To confirm this, each of the four SH-mutated OVAs, in which the original SS-forming sites (C11/73/120A, C30/73/120A, C73/120/367A, and C73/120/382A) were deleted, was constructed and expressed. The C11/73/120A and C30/73/120A mutants formed no SS form, in contrast to C73/120A as a control. Thus, we concluded that Cys30 participates in the correct folding of OVA, and that its SS bond (Cys11–Cys30) is transiently generated during the early folding stage to avoid misfolding, and then the native SS form of OVA is regenerated through SH–SS exchanges.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Physiological and genomic evidence of cysteine degradation and aerobic hydrogen sulfide production in freshwater bacteria

The sulfur-containing amino acid cysteine is abundant in the environment, including in freshwater lakes. Biological cysteine degradation can result in hydrogen sulfide (H 2 S), a toxic and ecologically relevant compound that is a central player in biogeochemical cycling in aquatic environments. Here, we investigated the ecological significance of cysteine in oxic freshwater, using isolated cultures, controlled experiments, and multiomics. We screened bacterial isolates enriched from natural lake water for their ability to produce H 2 S when provided cysteine. We identified 29 isolates (Bacteroidota, Proteobacteria, and Actinobacteria) that produced H 2 S. To understand the genomic and genetic basis for cysteine degradation and H 2 S production, we further characterized three isolates using whole-genome sequencing (using a combination of short-read and long-read sequencing) and tracked cysteine and H 2 S levels over their growth ranges: Stenotrophomonas maltophilia (Gammaproteobacteria), S. bentonitica (Gammaproteobacteria), and Chryseobacterium piscium (Bacteroidota). Cysteine decreased and H 2 S increased, and all three genomes had genes involved in cysteine degradation. Finally, to assess the presence of these organisms and genes in the environment, we surveyed a 5-year time series of metagenomic data from the same isolation source (Lake Mendota, Madison, WI, USA) and identified their presence throughout the time series. Overall, our study shows that diverse isolated bacterial strains can use cysteine and produce H 2 S under oxic conditions, and we show evidence using metagenomic data that this process may occur more broadly in natural freshwater lakes. Future considerations of sulfur cycling and biogeochemistry in oxic environments should account for H 2 S production from the degradation of organosulfur compounds.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Cysteine sensing by plasmons of silver nanocubes

Noble metal nanoparticles are considered to be valuable nanostructures in the field of sensors due to their spectral response sensitivity to small changes in the surrounding refractive index which enables them to detect a small amount of molecules. In this research, we use silver nanocubes of about 50 nm length to detect low concentrations of cysteine, a semi-essential amino acid. Following cysteine adsorption onto the nanocubes, a redshift in the plasmonic modes was observed, enabling the detection of cysteine down to 10 µM and high sensitivity of about 125 nm/RIU (refractive index units). Furthermore, we found that multilayer adsorption of cysteine leads to the stabilization of the silver nanocubes. The cysteine growth onto the nanocubes was also characterized by high-resolution transmission electron microscopy (HR-TEM). - Highlights: • Silver nanocubes (50 nm length) are used to detect low concentrations of cysteine. • A redshift in the plasmonic modes was observed following cysteine adsorption onto the nanocubes. • The cysteine growth onto the nanocubes is also characterized by TEM.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2016

Formation of Hg(II) tetrathiolate complexes with cysteine at neutral pH

Mercury(II) ions precipitate from aqueous cysteine (H 2 Cys) solutions containing H 2 Cys/Hg(II) mole ratio ≥ 2.0 as Hg( S -HCys) 2 . In absence of additional cysteine, the precipitate dissolves at pH ~12 with the [Hg( S , N -Cys) 2 ] 2- complex dominating. With excess cysteine (H 2 Cys/Hg(II) mole ratio ≥ 4.0), higher complexes form and the precipitate dissolves at lower pH values. Previously, we found that tetrathiolate [Hg( S -Cys) 4 ] 6- complexes form at pH = 11.0; in this work we extend the investigation to pH values of physiological interest. We examined two series of Hg(II)-cysteine solutions in which C Hg(II) varied between 8 – 9 mM and 80 – 100 mM, respectively, with H 2 Cys/Hg(II) mole ratios from 4 to ~20. The solutions were prepared in the pH range 7.1 – 8.8, at the pH at which the initial Hg( S -HCys) 2 precipitate dissolved. The variations in the Hg(II) speciation were followed by 199 Hg NMR, X-ray absorption and Raman spectroscopic techniques. Our results show that in the dilute solutions (C Hg(II) = 8 – 9 mM), mixtures of di-, tri- (major) and tetrathiolate complexes exist at moderate cysteine excess (C H2Cys ~ 0.16 M) at pH 7.1. In the more concentrated solutions (C Hg(II) = 80 – 100 mM) with high cysteine excess (C H2Cys > 0.9 M), tetrathiolate [Hg( S -cysteinate) 4 ] m-6 ( m = 0 – 4) complexes dominate in the pH range 7.3 – 7.8, with lower charge than for the [Hg( S -Cys) 4 ] 6- complex due to protonation of some ( m ) of the amino groups of the coordinated cysteine ligands. In conclusion, the results of this investigation could provide a key to the mechanism of biosorption and accumulation of Hg(II) ions in biological / environmental systems.

199Hg NMR↗
DOE OSTI2019

Synchrotron X-radiolysis of l -cysteine at the sulfur K-edge: Sulfurous products, experimental surprises, and dioxygen as an oxidoreductant

In-situ inventory of sulfurous products from the sulfur K-edge synchrotron X-radiolysis of L-cysteine, in solid-phase and anaerobic (pH 5) and air-saturated (pH 5, pH 7, and pH 9) solutions without and with 40% glycerol is reported. Sequential K-edge X-ray Absorption Spectroscopic (XAS) spectra were acquired. L-cysteine degraded systematically in the X-ray beam. Radiolytic products were inventoried by fits using the XAS spectra of sulfur model compounds. Solid L-cysteine declined to 92% fraction after a single K-edge XAS scan. After six scans 60 % remained, accompanied by 14 % cystine, 16 % thioether, 5.4 % of elemental sulfur, and smaller fractions of more highly oxidized products. In air-saturated pH 5 solution, 73 % of L-cysteine remained after ten scans, with 2 % cystine and 19 % elemental sulfur. Oxidation increased with 40% glycerol, yielding 67 %, 5 % and 23 % fractions, respectively, after ten-scans. Higher pH solutions exhibited less radiolytic chemistry. All the reactivity followed first-order kinetics. The anaerobic experiment displayed two reaction phases, with sharp changes in kinetics and radiolytic chemistry. Unexpectedly, the radiolytic oxidation of L-cysteine was increased in anaerobic solution. After ten-scans only 60% of the L-cysteine remained, along with 17 % cystine, 22 % elemental sulfur, and traces of more highly oxidized products. A new aerobic reaction cycle is hypothesized, wherein dissolved dioxygen captures radiolytic H• or e$-\atop{aq}$, enters HO 2 •/O 2 •-, reductively quenches cysteine thiyl radical, and cycles back to O 2 . This cycle is suggested to suppress the radiolytic production of cystine in aerobic solution.

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

Shining light on cysteine modification: connecting protein conformational dynamics to catalysis and regulation

We report that cysteine is a rare but functionally important amino acid that is often subject to covalent modification. Cysteine oxidation plays an important role in many human disease processes, and basal levels of cysteine oxidation are required for proper cellular function. Because reactive cysteine residues are typically ionized to the thiolate anion (Cys-S - ), their formation of a covalent bond alters the electrostatic and steric environment of the active site. X-ray-induced photo-oxidation to sulfenic acids (Cys-SOH) can recapitulate some aspects of the changes that occur under physiological conditions. Here we propose how site-specific cysteine photo-oxidation can be used to interrogate ensuing changes in protein structure and dynamics at atomic resolution. Although this powerful approach can connect cysteine covalent modification to global protein conformational changes and function, careful biochemical validation must accompany all such studies to exclude misleading artifacts. Lastly, new types of X-ray crystallography experiments and powerful computational methods are creating new opportunities to connect conformational dynamics to catalysis for the large class of systems that use covalently modified cysteine residues for catalysis or regulation.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Isolation, Characterization, and Depolymerization of L -Cysteine Substituted Eucalyptus Lignin

Lignin condensation reactions are hard to avoid or control during separation, which is a deterrent to lignin isolation and post-conversation, especially for the full utilization of lignocelluloses. Selective protection of β-aryl ether linkages in the isolation process is crucial to lignin valorization. Herein, a two-step acid/alkali separation method assisted with l-cysteine for eucalyptus lignin separation is developed, and the isolated L -cysteine lignins (LCLs) are comprehensively characterized by 2D NMR, 31 P NMR, thioacidolysis, etc. Compared to the two-step control treatment, a much higher β-O-4 content is preserved without reducing the separation efficiency assisted by L -cysteine, which is also significantly higher than alkali lignin and kraft lignin. The results of hydrogenolysis show that LCLs generate a much higher monomer yield than that of control sample. Structural analysis of LCLs suggests that lignin condensation reaction, to some extent, is suppressed by adding L -cysteine during the two-step acid/alkali separation. Further, mechanistic studies using dimeric model compound reveals that L -cysteine may be the α-carbon protective agent in the two-step separation. The role of L -cysteine in the two-step lignin isolation method provides novel insights to the selective fractionation of lignin from biomass, especially for the full valorization of lignocellulosic biomass.

09 BIOMASS FUELS↗
DOE OSTI2018

Structural basis of the cystein protease inhibitor Clonorchis sinensis Stefin-1

Highlights: • Clonorchis sinensis Stefin-1 has known as a host immune modulator. • The first crystal structure of CsStefin-1 was determined by the MAD method. • CsStefin-1 inhibited CsCF-8 cysteine protease activity. • A molecular docking model of CsStefin-1 and CsCF-8 was developed. • The CsStefin-1 was physically bound to CsCF-8 with an apparent K{sub D} value. Cystein protease plays a critical role as a virulence factor in the development and progression of various diseases. Cystatin is a superfamily of cysteine protease inhibitors that participates in various physiological and pathological processes. The cysteine protease inhibitor CsStein-1 isolated from Clonorchis sinensis belongs to the type 1 stefin of cystatins. This inhibitor regulates the activity and processing of CsCF (Cathepsin F of Clonorchis sienesis), which plays an important role in parasite nutrition and host-parasite interaction. CsStefin-1 has also been proposed as a host immune modulator and a participant in the mechanism associated with anti-inflammatory ability. Here, we report the first crystal structure of CsStefin-1 determined by the multi-wavelength anomalous diffraction (MAD) method to 2.3 Å. There are six molecules of CsStefin-1 per asymmetric unit, with a solvent content of 36.5%. The structure of CsStefin-1 is composed of twisted four-stranded antiparallel β-sheets, a central α-helix, and a short α-helix. We also demonstrate that CsStefin-1 binds to CsCF-8 cysteine protease and inhibits its activity. In addition, a molecular docking model of CsStefin-1 and CsCF-8 was developed using homology modeling based on their structures. The structural information regarding CsStefin-1 and molecular insight into its interaction with CsCF-8 are important to understanding their biological function and to design of inhibitors that modulate cysteine protease activity.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Purification and characterization of two cysteine proteinase inhibitors from silkworm, Bombyx mori

Highlights: • Two recombinant silkworm cysteine proteinase inhibitors were characterized. • The inhibitors showed different inhibitory activities toward cathepsin L. • Secondary structures of both inhibitors were analyzed at different temperatures. • The C-terminal tail was found to be necessary for BCPI inhibitory activity. • Arginine (R31) is important for the inhibitory activity in BCPI. Cysteine proteinase inhibitors from silkworm are selective inhibitors with low molecular weight and regulate cathepsin L-like cysteine proteinase activity, thus, affecting silkworm metamorphosis. In a previous study, two cysteine proteinase inhibitors, BCPI and BmCPI, were identified in the silkworm genome. To characterize these inhibitors, we expressed and purified them in an Escherichia coli system and analyzed their structure and inhibitory activity in vitro. Both inhibitors showed strong tolerance to high temperature. Their CD spectra revealed that their secondary structures could be recovered by a gradual decrease in temperature. Compared to BCPI, BmCPI exhibited weak inhibitory activity toward cathepsin L. BCPI activity was significantly decreased when its C-terminus was truncated, whereas BmCPI activity increased considerably when the C-terminus tail of BCPI was attached to BmCPI. Additionally, the inhibitory activity of BCPI was strongly reduced if R31 was mutated to A31. In summary, two cysteine proteinase inhibitors from silkworm were characterized in the present study, which facilitates an understanding of the interaction mechanism between cysteine proteinase and its inhibitors in the silkworm.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2015

Interactions between N-acetyl-L-cysteine protected CdTe quantum dots and doxorubicin through spectroscopic method

Highlights: • CdTe quantum dots with the diameter of 3–5 nm were synthesized in aqueous solution. • The modified CdTe quantum dots showed well fluorescence properties. • The interaction between the CdTe quantum dots and doxorubicin (DR) was investigated. - Abstract: N-acetyl-L-cysteine protected cadmium telluride quantum dots with a diameter of 3–5 nm were synthesized in aqueous solution. The interaction between N-acetyl-L-cysteine/cadmium telluride quantum dots and doxorubicin was investigated by ultraviolet–visible absorption and fluorescence spectroscopy at physiological conditions (pH 7.2, 37 °C). The results indicate that electron transfer has occurred between N-acetyl-L-cysteine/cadmium telluride quantum dots and doxorubicin under light illumination. The quantum dots react readily with doxorubicin to form a N-acetyl-L-cysteine/cadmium telluride-quantum dots/doxorubicin complex via electrostatic attraction between the −NH{sub 3}{sup +} moiety of doxorubicin and the −COO{sup −} moiety of N-acetyl-L-cysteine/cadmium telluride quantum dots. The interaction of N-acetyl-L-cysteine/cadmium telluride-quantum dots/doxorubicin complex with bovine serum albumin was studied as well, showing that the complex might induce the conformation change of bovine serum due to changes in microenvironment of bovine serum.

75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND↗