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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 181 records · Page 10
All sourcesNASA NTRSDOE OSTI
DOE OSTI2017

Green fluorescence protein-based content-mixing assay of SNARE-driven membrane fusion

Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins mediate intracellular membrane fusion by forming a ternary SNARE complex. A minimalist approach utilizing proteoliposomes with reconstituted SNARE proteins yielded a wealth of information pinpointing the molecular mechanism of SNARE-mediated fusion and its regulation by accessory proteins. Two important attributes of a membrane fusion are lipid-mixing and the formation of an aqueous passage between apposing membranes. These two attributes are typically observed by using various fluorescent dyes. Currently available in vitro assay systems for observing fusion pore opening have several weaknesses such as cargo-bleeding, incomplete removal of unencapsulated dyes, and inadequate information regarding the size of the fusion pore, limiting measurements of the final stage of membrane fusion. In the present study, we used a biotinylated green fluorescence protein and streptavidin conjugated with Dylight 594 (DyStrp) as a Föster resonance energy transfer (FRET) donor and acceptor, respectively. This FRET pair encapsulated in each v-vesicle containing synaptobrevin and t-vesicle containing a binary acceptor complex of syntaxin 1a and synaptosomal-associated protein 25 revealed the opening of a large fusion pore of more than 5 nm, without the unwanted signals from unencapsulated dyes or leakage. This system enabled determination of the stoichiometry of the merging vesicles because the FRET efficiency of the FRET pair depended on the molar ratio between dyes. Here, we report a robust and informative assay for SNARE-mediated fusion pore opening. - Highlights: • SNARE proteins drive membrane fusion and open a pore for cargo release. • Biotinylated GFP and DyStrp was used as the reporter pair of fusion pore opening. • Procedure for efficient SNARE reconstitution and reporter encapsulation was established. • The FRET pair reported opening of a large fusion pore bigger than 5 nm. • The assay was robust and provided information of stoichiometry of vesicle fusion.

60 APPLIED LIFE SCIENCES↗
NASA NTRS1987

Geomorphic evidence for ancient seas in west Deuteronilus Mensae, Mars-1: Regional geomorphology

The fretted terrain in west Deuteronilus Mensae consists of extensive cratered upland penninsulas or isolated plateaus cut by long, finger-like canyons typically 10 to 20 km wide and upwards of 300 km long. The longest of these canyons trend roughly north-south to north-northeast, which may reflect some local structural and/or topographic control. At least three geomorphic zones roughly parallel to the lowland/upland boundary, suggestive of increasing modification northward, can be recognized on the fretted region of the region. The southern-most zone (zone A) consists of sharply defined fretted terrain. The middle zone (zone B) consists of well defined fretted terrain in which the plateau surfaces appear smoother, with a somewhat darker and much less varied albedo surface than those of zone A. The northern-most zone (zone C) consists of rounded or softened fretted terrain. The zones were interpreted as surface exposures of successively lower stratigraphic units.

Parker, Timothy J.↗
NASA NTRS2023

Automated Verification of Programmable Logic Controller Programs Against Structured Natural Language Requirements

PLCverif is an actively developed project at CERN, enabling the formal verification of Programmable Logic Controller (PLC) programs in critical systems. In this paper, we present our work on improving the formal requirements specification experience in PLCverif through the use of natural language. To this end, we integrate NASA’s FRET, a formal requirement elicitation and authoring tool, into PLCverif. FRET is used to specify formal requirements in structured natural language, which automatically translates into temporal logic formulae. FRET’s output is then directly used by PLCverif for verification purposes. We discuss practical challenges that PLCverif users face when authoring requirements and the FRET features that help alleviate these problems. We present the new requirement formalization workflow and report our experience using it on two critical CERN case studies.

formal methods↗
DOE OSTI2013

Phase Behavior and Domain Size in Sphingomyelin-Containing Lipid Bilayers

Membrane raft size measurements are crucial to understanding the stability and functionality of rafts in cells. The challenge of accurately measuring raft size is evidenced by the disparate reports of domain sizes, which range from nanometers to microns for the ternary model membrane system sphingomyelin (SM)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/cholesterol (Chol). Using F rster resonance energy transfer (FRET) and differential scanning calorimetry (DSC), we established phase diagrams for porcine brain SM (bSM)/dioleoyl-sn-glycero-3-phosphocholine (DOPC)/Chol and bSM/POPC/Chol at 15 and 25 C. By combining two techniqueswith different spatial sensitivities, namely FRET and small-angle neutron scattering (SANS),we have significantly narrowed the uncertainty in domain size estimates for bSM/POPC/Chol mixtures. Compositional trends in FRET data revealed coexisting domains at 15 and 25 C for bothmixtures, while SANS measurements detected no domain formation for bSM/POPC/Chol. Together these results indicate that liquid domains in bSM/POPC/Chol are between 2 and 7 nmin radius at 25 C: that is, domains must be on the order of the 2 6 nmF rster distance of the FRET probes, but smaller than the ~7 nm minimum cluster size detectable with SANS. However, for palmitoyl SM (PSM)/POPC/Chol at a similar composition, SANS detected coexisting liquid domains. This increase in domain size upon replacing the natural SMcomponent (which consists of amixture of chain lengths) with synthetic PSM, suggests a role for SM chain length in modulating raft size in vivo.

Bio-SANS↗
DOE OSTI2013

The use of fluorescent intrabodies to detect endogenous gankyrin in living cancer cells

Expression of antibody fragments in mammalian cells (intrabodies) is used to probe the target protein or interfere with its biological function. We previously described the in vitro characterisation of a single-chain Fv (scFv) antibody fragment (F5) isolated from an intrabody library that binds to the oncoprotein gankyrin (GK) in solution. Here, we have isolated several other scFvs that interact with GK in the presence of F5 and tested whether they allow, when fused to fluorescent proteins, to detect by FRET endogenous GK in living cells. The binding of pairs of scFvs to GK was analysed by gel filtration and the ability of each scFv to mediate nuclear import/export of GK was determined. Binding between scFv-EGFP and RFP-labelled GK in living cells was detected by fluorescence lifetime imaging microscopy (FLIM). After co-transfection of two scFvs fused to EGFP and RFP, respectively, which form a tri-molecular complex with GK in vitro, FRET signal was measured. This system allowed us to observe that GK is monomeric and distributed throughout the cytoplasm and nucleus of several cancer cell lines. Our results show that pairs of fluorescently labelled intrabodies can be monitored by FLIM–FRET microscopy and that this technique allows the detection of lowly expressed endogenous proteins in single living cells. Highlights: ► Endogenous GK in living cells was targeted with pairs of fluorescently-tagged scFvs. ► Tri-molecular complexes containing two scFvs and one molecule GK were formed. ► GK was detected using fluorescence lifetime-based FRET imaging. ► GK is monomeric and homogeneously distributed in several cancer cell lines. ► This technique may have many applications in live-cell imaging of endogenous proteins.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2019

Effects of Donor–Acceptor Quantum Coherence and Non-Markovian Bath on the Distance Dependence of Resonance Energy Transfer

Accurate information on the distance dependence of resonance energy transfer (RET) is crucial for its utilization as a spectroscopic ruler of nanometer scale distances. In this regard, understanding the effects of donor-acceptor quantum coherence and non-Markovian bath, which become significant at short distances, has significant implications. The present work investigates this issue theoretically by comparing results from a theory of coherent RET (CRET) with a nonequilibrium version of Forster’s RET (FRET) theory, both accounting for non-Markovian bath effects. Even for a model where the donor-acceptor electronic coupling is of transition dipole interaction form, it is shown that the RET rate in general deviates from the inverse sixth power distance dependence as opposed to the prediction of the original FRET. On the one hand, it is shown that the donor-acceptor quantum coherence makes the distance dependence steeper than the sixth power, although detailed manner of enhancement is sensitive to specific values of parameters. On the other hand, the non-Markovian bath effects make the distance dependence more moderate than the sixth power for both CRET and nonueqilibrium FRET, because finite time scale of the bath causes the rate to be smaller than the prediction of original FRET. While these effects are demonstrated clearly in the population dynamics at sub-picosecond time scales, their contributions to the conventional RET efficiency are relatively minor. Furthermore, this indicates that the actual detection of such effects through conventional RET efficiency measurement requires either high precision or utilization of a donor with fast spontaneous decay rate of excitation.

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

Enhancement of Resonant Energy Transfer Due to an Evanescent Wave from the Metal

The high density of evanescent modes in the vicinity of a metal leads to enhancement of the near-field Förster resonant energy transfer (FRET) rate. In this paper, we present a classical approach to calculate the FRET rate based on the dyadic Green’s function of an arbitrary dielectric environment and consider the nonlocal limit of material permittivity in the case of the metallic half-space and thin film. In a dimer system, we find that the FRET rate is enhanced due to shared evanescent photon modes bridging a donor and an acceptor. Furthermore, a general expression for the FRET rate for multimer systems is derived. Finally, the presence of a dielectric environment and the path interference effect enhance the transfer rate, depending on the combination of distance and geometry.

36 MATERIALS SCIENCE↗
DOE OSTI2019

Sub-20 nm Core–Shell–Shell Nanoparticles for Bright Upconversion and Enhanced Förster Resonant Energy Transfer

Upconverting nanoparticles provide valuable benefits as optical probes for bioimaging and Förster resonant energy transfer (FRET) due to their high signal-to-noise ratio, photostability, and biocompatibility; yet, making nanoparticles small yields a significant decay in brightness due to increased surface quenching. Approaches to improve the brightness of UCNPs exist but often require increased nanoparticle size. Here we present a unique core-shell-shell nanoparticle architecture for small (sub-20 nm), bright upconversion with several key features: (1) maximal sensitizer concentration in the core for high near-infrared absorption, (2) efficient energy transfer between core and interior shell for strong emission, and (3) emitter localization near the nanoparticle surface for efficient FRET. This architecture consists of β-NaYbF 4 (core) @NaY 0.8-x Er x Gd 0.2 F 4 (interior shell) @NaY 0.8 Gd 0.2 F 4 (exterior shell), where sensitizer and emitter ions are partitioned into core and interior shell, respectively. Emitter concentration is varied (x = 1, 2, 5, 10, 20, 50, and 80%) to investigate influence on single particle brightness, upconversion quantum yield, decay lifetimes, and FRET coupling. We compare these seven samples with the field-standard core-shell architecture of β-NaY 0.58 Gd 0.2 Yb 0.2 Er 0.02 F 4 (core) @NaY 0.8 Gd 0.2 F 4 (shell), with sensitizer and emitter ions codoped in the core. At a single particle level, the core-shell-shell design was up to 2-fold brighter than the standard core-shell design. Further, by coupling a fluorescent dye to the surface of the two different architectures, we demonstrated up to 8-fold improved emission enhancement with the core-shell-shell compared to the core-shell design. Finally, we show how, given proper consideration for emitter concentration, we can design a unique nanoparticle architecture to yield comparable or improved brightness and FRET coupling within a small volume.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2012

Altering the Conjugation Pathway for Improved Performance of Benzobisoxazole-Based Polymer Guest Emitters in Polymer Light-Emitting Diodes

Benzobisoxazoles (BBOs) are known to increase the electron affinities and improve the electron transporting properties of materials containing them. However, BBO copolymers generally do not perform well as emissive guests in guest–host PLEDs due to inefficient Förster resonance energy transfer (FRET) between host and guest. The incomplete FRET results in a large amount of host emission and limits the potential efficiencies of the devices. In all previously reported BBO copolymers, the conjugation pathway was through the oxazole rings. Herein we report six new BBO copolymers with backbone connectivity directly on the central benzene ring, resulting in a conjugation pathway for the polymers that is perpendicular to the previously reported pathway. Guest–host PLEDs made using these polymers show that the new conjugation pathway improves FRET between the poly(N-vinylcarbazole) host and the BBO-containing polymer guest. Because of highly efficient FRET, no host emission is observed even at lower guest concentrations. The improved energy transfer results in devices with luminous efficiencies up to 3.1 Cd/A, a 3-fold improvement over previously reported BBO-based PLEDs. These results indicate that the conjugation pathway plays a critical role in designing emissive materials for guest–host PLEDs.

36 MATERIALS SCIENCE↗
DOE OSTI2014

Two-photon-excited fluorescence resonance energy transfer in an aqueous system of CdTe quantum dots and Rhodamine B

Two-photon excited fluorescence resonance energy transfer (FRET) between CdTe quantum dots with different emission peaks and Rhodamine B in aqueous solution are investigated both experimentally and theoretically. The photoluminescence and lifetime are measured using a time-resolved fluorescence test system. The two-photon excited FRET efficiency is found to increase as the degree of spectral overlap of the emission spectrum of CdTe and the absorption spectrum of Rhodamine B increases, which is due to the increase of Forster radius of the sample. Moreover, FRET efficiency increases when the ratio of acceptor/donor concentration increases. The two-photon excited FRET efficiency was found to reach 40%.

71 CLASSICAL AND QUANTUM MECHANICS, GENERAL PHYSIC↗
DOE OSTI2012

Imaging gene expression in real-time using aptamers

Signal transduction pathways are usually activated by external stimuli and are transient. The downstream changes such as transcription of the activated genes are also transient. Real-time detection of promoter activity is useful for understanding changes in gene expression, especially during cell differentiation and in development. A simple and reliable method for viewing gene expression in real time is not yet available. Reporter proteins such as fluorescent proteins and luciferase allow for non-invasive detection of the products of gene expression in living cells. However, current reporter systems do not provide for real-time imaging of promoter activity in living cells. This is because of the long time period after transcription required for fluorescent protein synthesis and maturation. We have developed an RNA reporter system for imaging in real-time to detect changes in promoter activity as they occur. The RNA reporter uses strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags), which can be expressed from a promoter of choice. The tobramycin, neomycin and PDC RNA aptamers have been utilized for this system and expressed in yeast from the GAL1 promoter. The IMAGEtag RNA kinetics were quantified by RT-qPCR. In yeast precultured in raffinose containing media the GAL1 promoter responded faster than in yeast precultured in glucose containing media. IMAGEtag RNA has relatively short half-life (5.5 min) in yeast. For imaging, the yeast cells are incubated with their ligands that are labeled with fluorescent dyes. To increase signal to noise, ligands have been separately conjugated with the FRET (Förster resonance energy transfer) pairs, Cy3 and Cy5. With these constructs, the transcribed aptamers can be imaged after activation of the promoter by galactose. FRET was confirmed with three different approaches, which were sensitized emission, acceptor photobleaching and donor lifetime by FLIM (fluorescence lifetime imaging microscopy). Real-time transcription was measured by FLIM-FRET, which was detected by the decrease in donor lifetime resulting from ligand binding to IMAGEtags that were newly synthesized from the activated GAL1 promoter. The FRET signal was specific for transcribed IMAGEtags.

Transcription↗
NASA NTRS1999

Fluorescence Studies of Protein Crystal Nucleation

Fluorescence can be used to study protein crystal nucleation through methods such as anisotropy, quenching, and resonance energy transfer (FRET), to follow pH and ionic strength changes, and follow events occurring at the growth interface. We have postulated, based upon a range of experimental evidence that the growth unit of tetragonal hen egg white lysozyme is an octamer. Several fluorescent derivatives of chicken egg white lysozyme have been prepared. The fluorescent probes lucifer yellow (LY), cascade blue, and 5-((2-aminoethyl)aminonapthalene-1-sulfonic acid (EDANS), have been covalently attached to ASP 101. All crystallize in the characteristic tetragonal form, indicating that the bound probes are likely laying within the active site cleft. Crystals of the LY and EDANS derivatives have been found to diffract to at least 1.7 A. A second group of derivatives is to the N-terminal amine group, and these do not crystallize as this site is part of the contact region between the adjacent 43 helix chains. However derivatives at these sites would not interfere with formation of the 43 helices in solution. Preliminary FRET studies have been carried out using N-terminal bound pyrene acetic acid (Ex 340 nm, Em 376 nm) lysozyme as a donor and LY (Ex -425 nm, Em 525 nm) labeled lysozyme as an acceptor. FRET data have been obtained at pH 4.6, 0.1 M NaAc buffer, at 5 and 7% NaCl, 4 C. The corresponding Csat values are 0.471 and 0.362 mg/ml (approximately 3.3 and approximately 2.5 x 10(exp -5) M respectively). The data at both salt concentrations show a consistent trend of decreasing fluorescence intensity of the donor species (PAA) with increasing total protein concentration. This decrease is more pronounced at 7% NaCl, consistent with the expected increased intermolecular interactions at higher salt concentrations reflected in the lower solubility. The calculated average distance between any two protein molecules at 5 x 10(exp -6) M is approximately 70nm, well beyond the range where any FRET can be expected. Results from these and ongoing studies will be presented.

Pusey, Marc L.↗
NASA NTRS2000

Fluorescence Studies of Protein Crystal Nucleation

We have postulated that, in the case of tetragonal chicken egg white lysozyme, crystal growth occurs by the addition of pre-critical nuclei sized n-mers that form in the bulk solution, and that the n-mer growth units were multiples of the tetrameric 4(sub 3) helical structure. These have the strongest intermolecular bonds in the crystal and are therefore likely to be the first species formed. High resolution AFM studies provide strong supporting evidence for this model, but the data also suggest that the actual species in solution may not be identical in structure to that found in the crystal. We are using fluorescence resonance energy transfer (FRET) to study the initial solution phase self-assembly process, using covalent fluorescent derivatives which crystallize in the characteristic P4(sub 3)2(sub 1)2(sub 1) space group. FRET studies are being carried out between the cascade blue (CB-lys, donor, Ex(sub max) 366 nm, Em 420 nm) and lucifer yellow (LY-lys, acceptor, Ex(sub max) 430 nm, Em 528 nm) asp101 derivatives. The estimated R(sub 0) for this probe pair, the distance where 50% of the donor energy is transferred to the acceptor, is approx. 1.2 nm, compared to 2.2 nm between the side chain carboxyls of adjacent asp101's in the crystalline 4(sub 3) helix. The short donor lifetime of 2.80 ns (chi(sup 2) = 0.644), coupled with the large average distances between the molecules (greater than or equal to 50 nm) in solution, ensure that any energy transfer observed is not due to random diffusive interactions. Lifetime data show that CB-lys has a single lifetime when it is the only species in solution. Similarly, LY-lys also exhibits a single lifetime of 4.63 ns (chi(sup 2) = 0.42) when alone in solution. Addition of LY-lys to CB-lys results in the appearance of a third lifetime component of 0.348ns for the CB-lys. The fractional intensities of the different species present can be used to estimate the distribution of monomer and n-mers in solution. The self-association process is a function of the protein concentration relative to the saturation concentration, and observing it in dilute solution (conc. less than or equal to 10(exp -5)M) requires that the experiments be performed under low solubility conditions, i.e., low temperatures and high salt concentrations. Data from preliminary steady state FRET studies with N-terminal bound pyrene acetic acid (PAA-lys, donor, Ex 340 nm, Em 376 nm) and asp101 LY-lys as an acceptor showed a consistent trend of decreasing donor fluorescence intensity with increasing total protein concentration. The FRET data have been obtained at pH 4.6, 0.1M NaAc buffer, at 5 and 7% NaCl, 4 C. The corresponding C(sub sat) values are 0.471 and 0.362 mg/ml (approx. 3.3 and approx. 2.5 x 10(exp -5)M respectively). The donor fluorescence decrease is more pronounced at7% NaCl, consistent with the expected increased intermolecular interactions at higher salt concentrations as reflected in the lower solubility. Results from these and other ongoing studies will be discussed in conjunction with an emerging model for how tetragonal lysozyme crystals nucleate and the relevance of that model to other proteins.

Pusey, Marc↗
NASA NTRS2004

Durability Assessment of Gamma Tial

Gamma TiAl was evaluated as a candidate alloy for low-pressure turbine blades in aeroengines. The durability of g-TiAl was studied by examining the effects of impact or fretting on its fatigue strength. Cast-to-size Ti-48Al-2Cr-2Nb was studied in impact testing with different size projectiles at various impact energies as the reference alloy and subsequently fatigue tested. Impacting degraded the residual fatigue life. However, under the ballistic impact conditions studied, it was concluded that the impacts expected in an aeroengine would not result in catastrophic damage, nor would the damage be severe enough to result in a fatigue failure under the anticipated design loads. In addition, other gamma alloys were investigated including another cast-to-size alloy, several cast and machined specimens, and a forged alloy. Within this Ti-48-2-2 family of alloys aluminum content was also varied. The cracking patterns as a result of impacting were documented and correlated with impact variables. The cracking type and severity was reasonably predicted using finite element models. Mean stress affects were also studied on impact-damaged fatigue samples. The fatigue strength was accurately predicted based on the flaw size using a threshold-based, fracture mechanics approach. To study the effects of wear due to potential applications in a blade-disk dovetail arrangement, the machined Ti-47-2-2 alloy was fretted against In-718 using pin-on-disk experiments. Wear mechanisms were documented and compared to those of Ti-6Al-4V. A few fatigue samples were also fretted and subsequently fatigue tested. It was found that under the conditions studied, the fretting was not severe enough to affect the fatigue strength of g-TiAl.

Draper, Susan L.↗
NASA NTRSDate not supplied

Authoring, Analyzing, and Monitoring Requirements for a Lift-Plus-Cruise Aircraft

Requirements specification and analysis is widely applied to ensure the correctness of industrial systems in safety critical domains. Requirements are often initially written in natural language, which is highly ambiguous, and as a second step transformed into a language with rigorous semantics for formal analysis. [Question/problem] In this paper, we report on our experience in requirements creation and analysis, as well as run-time monitor generation using the Formal Requirement Elicitation Tool (FRET), on an industrial case study for a Lift-Plus-Cruise concept aircraft. [Principal ideas/results] We study the creation of requirements directly in the structured language of FRET without a prior definition of the same requirements in natural language. We focus on requirements describing state machines and discuss the challenges that we faced, in terms of creating requirements and generating monitors. We demonstrate how realizability, i.e., checking whether a requirements specification can be implemented, is crucial for understanding temporal interdependencies among requirements. [Contribution] Our study is the first complete attempt at using FRET to create industrial, realizable requirements and generate run-time monitors. Insight from lessons learned was materialized into new features in the FRET and JKind analysis frameworks.

Requirements engineering↗
NASA NTRSDate not supplied

Authoring, Analyzing, and Monitoring Requirements for a Lift-Plus-Cruise Aircraft

Requirements specification and analysis is widely applied to ensure the correctness of industrial systems in safety critical domains. Requirements are often initially written in natural language, which is highly ambiguous, and as a second step transformed into a language with rigorous semantics for formal analysis. In this paper, we report on our experience in requirements creation and analysis, as well as run-time monitor generation using the Formal Requirement Elicitation Tool (FRET), on an industrial case study for a Lift-Plus-Cruise concept aircraft. We study the creation of requirements directly in the structured language of FRET without a prior definition of the same requirements in natural language. We focus on requirements describing state machines and discuss the challenges that we faced, in terms of creating requirements and generating monitors. We demonstrate how realizability, i.e., checking whether a requirements specification can be implemented, is crucial for understanding temporal interdependencies among requirements. Our study is the first complete attempt at using FRET to create industrial, realizable requirements and generate run-time monitors. Insight from lessons learned was materialized into new features in the FRET and JKind analysis frameworks.

Requirements engineering↗
DOE OSTI2016

Assessment of wear coefficients of nuclear zirconium claddings without and with pre-oxidation

In the cores of pressurized water nuclear reactors, water-flow induced vibration is known to cause claddings on the fuel rods to rub against their supporting grids. Such grid-to-rod-fretting (GTRF) may lead to fretting wear-through and the leakage of radioactive species. The surfaces of actual zirconium alloy claddings in a reactor are inevitably oxidized in the high-temperature pressurized water, and some claddings are even pre-oxidized. As a result, the wear process of the surface oxide film is expected to be quite different from the zirconium alloy substrate. In this paper, we attempt to measure the wear coefficients of zirconium claddings without and with pre-oxidation rubbing against grid samples using a bench-scale fretting tribometer. Results suggest that the volumetric wear coefficient of the pre-oxidized cladding is 50 to 200 times lower than that of the untreated cladding. In terms of the linear rate of wear depth, the pre-oxidized alloy wears about 15 times more slowly than the untreated cladding. Finally, fitted with the experimentally-determined wear rates, a stage-wise GTRF engineering wear model demonstrates good agreement with in-reactor experience in predicting the trend of cladding lives.

21 SPECIFIC NUCLEAR REACTORS AND ASSOCIATED PLANTS↗
DOE OSTI2011

Dynamic behavior of small heat shock protein inhibition on amyloid fibrillization of a small peptide (SSTSAA) from RNase A

Highlights: Black-Right-Pointing-Pointer Mechanism of small heat shock protein inhibition on fibril formation was studied. Black-Right-Pointing-Pointer Peptide SSTSAA with modified ends was used for amyloid fibril formation. Black-Right-Pointing-Pointer FRET signal was followed during the fibril formation. Black-Right-Pointing-Pointer Mj HSP16.5 inhibits fibril formation when introduced in the lag phase. Black-Right-Pointing-Pointer Mj HSP16.5 slows down fibril formation when introduced after the lag phase. -- Abstract: Small heat shock proteins, a class of molecular chaperones, are reported to inhibit amyloid fibril formation in vitro, while the mechanism of inhibition remains unknown. In the present study, we investigated the mechanism by which Mj HSP16.5 inhibits amyloid fibril formation of a small peptide (SSTSAA) from RNase A. A model peptide (dansyl-SSTSAA-W) was designed by introducing a pair of fluorescence resonance energy transfer (FRET) probes into the peptide, allowing for the monitoring of fibril formation by this experimental model. Mj HSP16.5 completely inhibited fibril formation of the model peptide at a molar ratio of 1:120. The dynamic process of fibril formation, revealed by FRET, circular dichroism, and electron microscopy, showed a lag phase of about 2 h followed by a fast growth period. The effect of Mj HSP16.5 on amyloid fibril formation was investigated by adding it into the incubation solution during different growth phases. Adding Mj HSP16.5 to the incubating peptide before or during the lag phase completely inhibited fibril formation. However, introducing Mj HSP16.5 after the lag phase only slowed down the fibril formation process by adhering to the already formed fibrils. These findings provide insight into the inhibitory roles of small heat shock proteins on amyloid fibril formation at the molecular level.

60 APPLIED LIFE SCIENCES↗