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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 217 records · Page 12
All sourcesNASA NTRSDOE OSTI
NASA NTRS1996

Effects of Purification on the Crystallization of Lysozyme

We have additionally purified a commercial lysozyme preparation by cation exchange chromatography, followed by recrystallization. This material is 99.96% pure with respect to macromolecular impurities. At basic pH, the purified lysozyme gave only tetragonal crystals at 20 C. Protein used directly from the bottle, prepared by dialysis against distilled water, or which did not bind to the cation exchange column had considerably altered crystallization behavior. Lysozyme which did not bind to the cation exchange column was subsequently purified by size exclusion chromatography. This material gave predominately bundles of rod-shaped crystals with some small tetragonal crystals at lower pHs. The origin of the bundled rod habit was postulated to be a thermally dependent tetragonal- orthorhombic change in the protein structure. This was subsequently ruled out on the basis of crystallization behavior and growth rate experiments. This suggests that heterogeneous forms of lysozyme may be responsible. These results demonstrate three classes of impurities: (1) small molecules, which may be removed by dialysis; (2) macromolecules, which are removable by chromatographic techniques; and (3) heterogeneous forms of the protein, which can be removed in this case by cation exchange chromatography. Of these, heterogeneous forms of the lysozyme apparently have the greatest affect on its crystallization behavior.

Ewing, Felecia L.↗
NASA NTRS1973

Studies of a Halophilic NADH Dehydrogenase. 1: Purification and Properties of the Enzyme

An NADH dehydrogenase obtained from an extremely halophilic bacterium was purified 570-fold by a combination of gel filtration, chromatography on hydroxyapatite, and ion-exchange chromatography on QAE-Sephadex. The purified enzyme appeared to be FAD-linked and bad an apparent molecular weight of 64000. Even though enzyme activity was stimulated by NaCl, considerable activity (430 % of the maximum activity observed in the presence of 2.5 M NaCl) was observed in the absence of added NaCl. The enzyme was unstable when incubated in solutions of low ionic strength. The presence of NADH enhanced the stability of the enzyme.

Hochstein, Lawrence I.↗
NASA NTRS1992

Purification and Properties of an ATPase from Sulfolobus solfataricus

A sulfite-activated ATPase isolated from Sulfolobus solfataricus had a relative molecular mass of 370,000. It was composed of three subunits whose relative molecular masses were 63,000, 48,000, and 24,000. The enzyme was inhibited by the vacuolar ATPase inhibitors nitrate and N-ethylmaleimide; 4-chloro-7-nitrobenzo-furazan (NBD-Cl) was also inhibitory. N-Ethylmaleimide was predominately bound to the largest subunit while NBD-CL was bound to both subunits. ATPase activity was inhibited by low concentrations of p-chloromercuri-phenyl sulfonate and the inhibition was reversed by cysteine which suggested that thiol groups were essential for activity. While the ATPase from S. solfataricus shared several properties with the ATPase from S. acidocaldarius there were significant differences. The latter enzyme was activated by sulfate and chloride and was unaffected by N-ethylmaleimide, whereas the S. solfataricus ATPase was inhibited by these anions as well as N-ethyimaleimide. These differences as well as differences that occur in other vacuolar-like ATPases isolated from the methanogenic and the extremely halophilic bacteria suggest the existence of several types of archaeal ATPases, none of which have been demonstrated to synthesize ATP.

Hochstein, Lawrence I.↗
NASA NTRS1999

The Design and Development of a Regenerative Separatory Column Using Calixarenes as a Polymeric Backbone for the Purification of Water from Urine

The objective of this research project is to design calixarenes, cup-shaped molecules, with the specific binding sites to the sodium chloride and nitrogen containing components of urine, such as urea and uric acid, in urine. The following partition of the research accomplishes this objective: (1) functionalization of calixarene, (2) development of a calixarene based medium for the separatory process, (3) design of the column regeneration protocol. Work was also accomplished in the area of temperature sensitive paint (TSP). Research was undertaken to design a TSP with insulating propertites. An important part of this research project is to discover the thermal conductivity of polymers for TSP.

Polk, M.↗
NASA NTRS1998

Heterogeneity Determination and Purification of Commercial Hen Egg-White Lysozyme

Hen egg-white lysozyme (HEWL) is widely used as a model protein, although its purity has not been adequately characterized by modern biochemical techniques. We have identified and quantified the protein heterogeneities in three commercial HEWL preparations by sodium dodecyl sulfate polyacrylamide gel electrophoresis with enhanced silver staining, reversed-phase fast protein liquid chromatography (FPLC) and immunoblotting with comparison to authentic protein standards. Depending on the source, the contaminating proteins totalled 1-6%(w/w) and consisted of ovotransferrin, ovalbumin, HEWL dimers, and polypeptides with approximate M(sub r) of 39 and 18 kDa. Furthermore, we have obtained gram quantities of electrophoretically homogeneous [> 99.9%(w/w)] HEWL by single-step semi-preparative scale cation-exchange FPLC with a yield of about 50%. Parallel studies of crystal growth kinetics, salt repartitioning and crystal perfection with this highly purified material showed fourfold increases in the growth-step velocities and significant enhancement in the structural homogeneity of HEWL crystals.

Thomas, B. R.↗
NASA NTRS1998

Purification and Crystal Growth of Lead Iodide by Physical Vapor Transport Method

Lead iodide (PbI2) is a layered compound semiconductor being developed as room temperature x- and gamma-ray detector. Compared to the more studied material, mercuric iodide, PbI2 has a higher melting temperature and no phase transition until liquid phase which are indications of better mechanical properties. In this study, the source material was purified by the zone-refining process, and the purest section was extracted from center of the the zone-refined ingot to be grown by physical vapor transport (PVT) method. The zone-refined material and as-grown crystals were characterized by optical microscopy and differential scanning calorimetry (DSC) to reveal the surface morphology, purity and stoichiometry. The results shows that both materials are near-stoichiometric composition, with the purity of the as-grown crystals higher than zone-refined materials. The resistivity of the as-grown crystal (10" Omega-cm) was derived from current-voltage (I-V) measurement, and is 10 times higher than the zone-refined materials. Detail results will be presented and discussed.

Wright, G. W.↗
NASA NTRS2000

Purification of Restriction Endonuclease EcoRII and its Co-Crystallization

Restriction endonuclease EcoRII (EcoRII) is a homodimeric DNA-binding protein. It belongs to the type II family of restriction-modification enzymes (subclass IIe). EcoRII recognizes the nucleotide sequence 5'-CCWGG (W=A or T) and cleaves the phosphodiester bond preceding the first cytosine. Methylation at C5 of the second cytosine inhibits cleavage. The enzyme has a unique ability to search for the presence of two substrate sites before cleavage. To the best of our knowledge no other subclass IIe restriction endonuclease has been crystallized yet, without or with a DNA-substrate. We have recently grown and characterized the crystals of this enzyme (1) Here we report on the result of co-crystallization experiments of EcoRII with an 11 b.p. oligonucleotide substrate. The dissociation constant (Kd) EcoRII: 11 b.p. was determined earlier (unpublished results). The needle-like crystals of oligonucleotide-EcoRII protein complex were obtained with this substrate by the technique of vapor diffusion hanging drops. The crystals obtained were washed and dissolved in an aliquot of 10 mM Tris-HCl buffer, pH=7.5. Running a portion of this solution on the SDS-get indicated the presence of endonuclease in the solution. A UV-spectrophotometric test of a second portion confirmed the presence of DNA. We are now working on improvement of the DNA-EcoRII protein crystals. Results obtained from these and ongoing efforts will be reported.

Karpova, E. A.↗
NASA NTRS2001

The Purification and Characterization of Superoxide Dismutase from Chloroflexus aurantiacus and the Effects of UV Radiation on the Activity of SOD and Catalase in Hydrothermal Mats of Yellowstone National Park

Chloroflexus aurantiacus is a thermotolerant anoxygenic green phototrophic bacterium that is prominent in alkaline hot springs at temperatures between 52 and 60 C. This species often grows in the hyperoxic environment beneath cyanobacterial mats at higher temperatures up to 70 - 72 C. Cf. aurantiacus is an evolutionarily important organism since it is in the earliest branch of the eubacteria that are capable of photosynthesis and many of its characteristics can be found in other diverse groups of phototrophic bacteria. Additional information is contained in the original extended abstract.

Lancaster, Vanessa↗
NASA NTRS1994

Water Purification

The Vision Catalyst Purifier employs the basic technology developed by NASA to purify water aboard the Apollo spacecraft. However, it also uses an "erosion" technique. The purifier kills bacteria, viruses, and algae by "catalytic corrosion." A cartridge contains a silver-impregnated alumina bed with a large surface area. The catalyst bed converts oxygen in a pool of water to its most oxidative state, killing over 99 percent of the bacteria within five seconds. The cartridge also releases into the pool low levels of ionic silver and copper through a controlled process of erosion. Because the water becomes electrochemically active, no electricity is required.

Source record↗
NASA NTRS1992

Water Purification Systems

A water purification/recycling system developed by Photo-Catalytics, Inc. (PCI) for NASA is commercially available. The system cleanses and recycles water, using a "photo-catalysis" process in which light or radiant energy sparks a chemical reaction. Chemically stable semiconductor powders are added to organically polluted water. The powder absorbs ultraviolet light, and pollutants are oxidized and converted to carbon dioxide. Potential markets for the system include research and pharmaceutical manufacturing applications, as well as microchip manufacture and wastewater cleansing.

Source record↗
NASA NTRS2002

Solutal Convection Around Growing Protein Crystal and Diffusional Purification in Space

This work theoretically addressed two subjects: 1) onset of convection, 2) distribution of impurities. Onset of convection was considered analytically and numerically. Crystal growth was characterized by slow surface incorporation kinetics, i.e. growth kinetic coefficient beta (cm/s) small as compared to the typical bulk diffusion rate, D(sub 1)/h, where D(sub 1) is diffusivity of major crystallizing protein and h is the crystal size. Scaling type analysis predicted two laws on how the convection rate, v, essentially the Peclet number, Pe exactly equal to vh/D(sub 1), depends on dimensionless kinetic coefficient a exactly equal to beta h/D(sub 1). Namely: Pe = C(sub 2/5)(aRa(sup 2/5)) and Pe = C(sub 1) aRa. Here, Reynolds number Ra = rho(sub 1)(sup 0)gh(sup 3)(rho(sub p) - rho(sub w))/rho(sup p)rho(sub 1)vD(sub 1), v being solution viscosity. The constants C(sub 2/5), exactly equal to 0.28 and C(sub 1) exactly equal to 10(exp -2) found from the full scale computer simulation for a cylindrical crystal inside big cylindrical vessel. The linear boundary conditions connecting protein and impurity concentration at the interface with the flux to/from the interface was applied. No-slip condition for Navier-Shocker equations was employed. With these conditions, flow and concentration distributions were calculated. Validity of the Pe(Ra) dependencies follows for wide range of parameters for which numerical calculations have been accomplished and presented by various points.

Chernov, A. A.↗
NASA NTRS2003

Carbon Nanotube Purification and Functionalization

Carbon nanotubes have the potential to significantly enhance the mechanical, thermal, and electrical properties of polymers. However, dispersion of carbon nanotubes in a polymer matrix is hindered by the electrostatic forces that cause them to agglomerate. Chemical modification of the nanotubes is necessary to minimize these electrostatic forces and promote adhesion between the nanotubes and the polymer matrix. In a collaborative research program between Clark Atlanta University, Rice University, and NASA Glenn Research Center several approaches are being explored to chemically modify carbon nanotubes. The results of this research will be presented.

Lebron, Marisabel↗
NASA NTRS2003

Purification, Crystallization, and Preliminary X-ray Analysis of Native Canavalin

The protein canavalin is a 7S vicilin, from the Jack Bean, Canavalis ensfomis. Canavalin is described as a seed storage protein, an energy source for a developing seed, as no other known activity or function has been found. The protein was first isolated and crystallized by Sumner and Howell (J. Biol. Chem. 113, 607-610, 1936). Canavalin spontaneously crystallizes after proteolytic cleavage at neutral pH, which removes residues 1-46, 224-245, and 325-330 and produces peptides of approximately 25, 13, and 12 kDa. Preliminary gel filtration experiments indicated the presence of nucleic acid with the uncleaved protein. We developed a dual column procedure, ion exchange followed by hydroxy apatite chromatography, that effectively removes the nucleic acid and yields an essentially pure uncut canavalin preparation with an OD 280/260 ratio of approximately 1.9-2.0. Standard crystallization screens using this material gave a number of positive results having a common requirement for alcohols and Mg(2+) ion, with crystals typically appearing within a day or less. Optimization experiments to date have shown that we can obtain crystals from pH 6.5 to pH 8.2, using MPD from 5 to 20% and 0.05 to 0.2M Mg(2+) (sulfate or acetate). The crystals are of space group P2(sub 1)2(sub 1)2(sub 1), unit cell dimensions, and a complete data set to 1.5 Angstroms, resolution has now been collected at a synchrotron source. Most importantly, the crystals are not twinned, a persistent problem with the most commonly obtained rhombohedral form of proteolytically cleaved canavalin.

Pusey, Marc↗
NASA NTRS2004

Solutal Convection Around Growing Protein Crystal and Diffusional Purification in Space

At least some protein crystals were found to preferentially trap microheterogeneous impurities. The latter are, for example, dimmer molecules of the crystallizing proteines (e.g. ferritin, lysozyme), or the regular molecules on which surfaces small molecules or ions are adsorbed (e.g. acetilated lysozyme) and modi@ molecular charge. Impurities may induce lattice defects and deteriorate structural resolution. Distribution of impurities between mother solution and gorwing crystal is defined by two interrelated distribution coefficients: kappa = rho(sup c, sub 2) and K = (rho(sup c, sub 2)/rho(sup c, sub 1)/rho(sub 2)/rho(sub 1). Here, rho(sub 2), rho(sub 1) and rho(sup c, sub 2) are densities of impurity (2) and regular protein (1) in solution at the growing interface and within the crystal ("c"). For the microheterogeneous impurities studied, K approx. = 2 - 4, so that kappa approx. - 10(exp 2) - 10(exp 3), since K = kappa (rho(sub 1)/rho(sup c, sub 1) and protein solubility ratio rho(sub 1)/rho(sub=p c, sub 2) much less than 1. Therefore, a crystal growing in absence of convection purifies mother solution around itself, grows cleaner and, probably, more perfect. If convection is present, the solution flow permanently brings new impurities to the crystal. This work theoretically addressed two subjects: 1) onset of convection, 2) distribution of impurities.

Lee, Chun P.↗
NASA NTRS1999

A novel beta-glucosidase from the cell wall of maize (Zea mays L.): rapid purification and partial characterization

Plants have a variety of glycosidic conjugates of hormones, defense compounds, and other molecules that are hydrolyzed by beta-glucosidases (beta-D-glucoside glucohydrolases, E.C. 3.2.1.21). Workers have reported several beta-glucosidases from maize (Zea mays L.; Poaceae), but have localized them mostly by indirect means. We have purified and partly characterized a 58-Ku beta-glucosidase from maize, which we conclude from a partial sequence analysis, from kinetic data, and from its localization is not identical to any of those already reported. A monoclonal antibody, mWP 19, binds this enzyme, and localizes it in the cell walls of maize coleoptiles. An earlier report showed that mWP19 inhibits peroxidase activity in crude cell wall extracts and can immunoprecipitate peroxidase activity from these extracts, yet purified preparations of the 58 Ku protein had little or no peroxidase activity. The level of sequence similarity between beta-glucosidases and peroxidases makes it unlikely that these enzymes share epitopes in common. Contrary to a previous conclusion, these results suggest that the enzyme recognized by mWP19 is not a peroxidase, but there is a wall peroxidase closely associated with the 58 Ku beta-glucosidase in crude preparations. Other workers also have co-purified distinct proteins with beta-glucosidases. We found no significant charge in the level of immunodetectable beta-glucosidase in mesocotyls or coleoptiles that precedes the red light-induced changes in the growth rate of these tissues.

Non-NASA Center↗
NASA NTRS1986

The purification and properties of a cd-cytochrome nitrite reductase from Paracoccus halodenitrificans

Paracoccus halodenitrificans, grown anaerobically in the presence of nitrite, contained membrane and cytoplasmic nitrite reductases. When assayed in the presence of phenazine methosulfate and ascorbate, the membrane-bound enzyme produced nitrous oxide whereas the cytoplasmic enzyme produced nitric oxide. When both enzymes were assayed in the presence of methyl viologen and dithionite, the cytoplasmic enzyme produced ammonia. Following solubilization, the membrane-bound enzyme behaved like the cytoplasmic enzyme, producing nitric oxide in the presence of phenazine methosulfate and ascorbate, and ammonia when assayed in the presence of methyl viologen and dithionite. The cytoplasmic and membrane-bound enzymes were purified to essentially the same specific activity. Only a single nitrite-reductase activity was detected on electrophoretic gels and the electrophoretic behavior of both enzymes suggested they were identical. The spectral properties of both enzymes suggested they were cd-type cytochromes. These data suggest that the products of nitrite reduction by the cd-cytochrome nitrite reductase are determined by the location of the enzyme and the redox potential of the electron donor.

NASA Center ARC↗
NASA NTRS1995

Simultaneous concentration and purification through gradient deformation chromatography

Mobile-phase additives, commonly used to modulate absorbate retention in gradient elution chromatography, are usually assumed to be either linearly retained or unretained. Previous theoretical work from our laboratory has shown that these modulators, such as salts in ion-exchange and hydrophobic interaction chromatography and organic modifiers in reversed-phase chromatography, can absorb nonlinearly, giving rise to gradient deformation. Consequently, adsorbate peaks that elute in the vicinity of the head of the deformed gradient may exhibit unusual shapes, form shoulders, and/or be concentrated. These effects for a reversed-phase sorbent with aqueous acetonitrile (ACN) as the modulator are verified experimentally. Gradient deformation is demonstrated experimentally and agrees with simulations based on ACN isotherm parameters that are independently determined from batch equilibrium studies using the layer model. Unusual absorbate peak shapes were found experimentally for single-component injections of phenylalanine, similar to those calculated by the simulations. A binary mixture of tryptophan and phenylalanine is used to demonstrate simultaneous concentration and separation, again in agreement with simulations. The possibility of gradient deformation in ion-exchange and hydrophobic interaction chromatography is discussed.

NASA Discipline Number 93-10↗
NASA NTRS1991

Partial purification and characterization of a Ca(2+)-dependent protein kinase from pea nuclei

Almost all the Ca(2+)-dependent protein kinase activity in nuclei purified from etiolated pea (Pisum sativum, L.) plumules is present in a single enzyme that can be extracted from chromatin by 0.3 molar NaCl. This protein kinase can be further purified 80,000-fold by salt fractionation and high performance liquid chromatography, after which it has a high specific activity of about 100 picomoles per minute per microgram in the presence of Ca2+ and reaches half-maximal activation at about 3 x 10(-7) molar free Ca2+, without calmodulin. It is a monomer with a molecular weight near 90,000. It can efficiently use histone III-S, ribosomal S6 protein, and casein as artificial substrates, but it phosphorylates phosvitin only weakly. Its Ca(2+)-dependent kinase activity is half-maximally inhibited by 0.1 millimolar chlorpromazine, by 35 nanomolar K-252a and by 7 nanomolar staurosporine. It is insensitive to sphingosine, an inhibitor of protein kinase C, and to basic polypeptides that block other Ca(2+)-dependent protein kinases. It is not stimulated by exogenous phospholipids or fatty acids. In intact isolated pea nuclei it preferentially phosphorylates several chromatin-associated proteins, with the most phosphorylated protein band being near the same molecular weight (43,000) as a nuclear protein substrate whose phosphorylation has been reported to be stimulated by phytochrome in a calcium-dependent fashion.

NASA Discipline Number 40-50↗