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At least 649 records · Page 36
All sourcesNASA NTRSDOE OSTI
NASA NTRS1987

Mixing Valve for Protein-Crystal Growth

Apparatus for growing protein and other crystals holds reactants in separate chambers, then allows them to combine at proper time. Openings in cylindrical rotary valve connect chambers to each other selectively through internal passages. O-ring and strip seals prevent leakage over valve surface. Caps on chambers seal them tightly but are removable for addition of reactants or withdrawal of reaction products. Made from various chemically inert materials, and chambers designed with capacities ranging from milliliters to liters. Easily automated.

Carter, Daniel C.↗
NASA NTRS1987

Protein crystal growth in low gravity

The solubility and growth mechanism of canavalin were studied, and the applicability of the Schlieren technique to protein crystal growth was investigated. Canavalin which may be crystallized from a basic solution by the addition of hydrogen (H+) ions was shown to have normal solubility characteristics over the range of temperatures (5 to 25 C) and pH (5 to 7.5) studied. The solubility data combined with growth rate data gathered from the seeded growth of canavalin crystals indicated that the growth mechanism at high supersaturation ratios (>1.28) is screw dislocation like. A Schlieren apparatus was constructed and flow patterns were observed in Rochelle salt (sodium potassium tartrate), lysozyme, and canavalin. The critical parameters were identified as the change in density with concentration (dp/dc) and the change in index of refraction with concentration (dn/dc). Some of these values were measured for the materials listed.

Feigelson, Robert S.↗
NASA NTRS1987

Model for determining vapor equilibrium rates in the hanging drop method for protein crystal growth

An engineering analysis of the rate of evaporation of solvent in the hanging drop method of protein crystal growth is presented. Results are applied to 18 drop and well arrangements commonly encountered in the laboratory. The chemical nature of the salt, drop size and shape, drop concentration, well size, well concentration, and temperature are taken into account. The rate of evaporation increases with temperature, drop size, and the salt concentration difference between the drop and the well. The evaporation in this model possesses no unique half-life. Once the salt in the drop achieves 80 percent of its final concentration, further evaporation suffers from the law of diminishing returns.

Baird, James K.↗
NASA NTRS1986

Protein crystal growth - Growth kinetics for tetragonal lysozyme crystals

Results are reported from theoretical and experimental studies of the growth rate of lysozyme as a function of diffusion in earth-gravity conditions. The investigations were carried out to form a comparison database for future studies of protein crystal growth in the microgravity environment of space. A diffusion-convection model is presented for predicting crystal growth rates in the presence of solutal concentration gradients. Techniques used to grow and monitor the growth of hen egg white lysozyme are detailed. The model calculations and experiment data are employed to discuss the effects of transport and interfacial kinetics in the growth of the crystals, which gradually diminished the free energy in the growth solution. Density gradient-driven convection, caused by presence of the gravity field, was a limiting factor in the growth rate.

Pusey, M. L.↗
NASA NTRS1988

Optical monitoring of protein crystal growth

The possibility of using various optical techniques for detecting the onset of nucleation in protein crystal growth was investigated. Direct microscopy, general metrologic techniques, light scattering, ultraviolet absorption, and interferometry are addressed along with techniques for determining pH value. The necessity for collecting basic data on the optical properties of the growth solution as a prerequisite to the evaluation of monitoring techniques is pointed out.

Choudry, A.↗
NASA NTRS1987

Evaporation kinetics in the hanging drop method of protein crystal growth

An engineering analysis of the rate of evaporation of solvent in the hanging drop method of protein crystal growth is presented; these results are applied to 18 different drop and well arrangements commonly encountered in the laboratory, taking into account the chemical nature of the salt, the drop size and shape, the drop concentration, the well size, the well concentration, and the temperature. It is found that the rate of evaporation increases with temperature, drop size, and with the salt concentration difference between the drop and the well. The evaporation possesses no unique half-life. Once the salt in the drop achieves about 80 percent of its final concentration, further evaporation suffers from the law of diminishing returns.

Baird, James K.↗
NASA NTRS1987

Continuous flow electrophoretic separation of proteins and cells from mammalian tissues

This paper describes an apparatus for continuous flow electrophoresis (CFE), designed to separate macromolecules and cells at conditions of microgravity. In this CFE, buffer flows upward in a 120-cm long flow chamber, which is 16-cm wide x 3.0-mm thick in the microgravity version (and 6-cm wide x 1.5-mm thick in the unit-gravity laboratory version). Ovalbumin and rat serum albumin were separated in space (flight STS-4) with the same resolution of the two proteins achieved at 25 percent total w/v concentration that was obtained in the laboratory at 0.2 percent w/v concentration. Rat anterior pituitary cells, cultured human embryonic kidney cells, and canine Langerhans cells were separated into subpopulations (flight STS-8) more effectively than in unit gravity, with comparable resolution having been achieved at 100 times the concentration possible on earth.

Hymer, W. C.↗
NASA NTRS1988

A simple apparatus for controlling nucleation and size in protein crystal growth

A simple device is described for controlling vapor equilibrium in macromolecular crystallization as applied to the protein crystal growth technique commonly referred to as the 'hanging drop' method. Crystal growth experiments with hen egg white lysozyme have demonstrated control of the nucleation rate. Nucleation rate and final crystal size have been found to be highly dependent upon the rate at which critical supersaturation is approached. Slower approaches show a marked decrease in the nucleation rate and an increase in crystal size.

Gernert, Kim M.↗
NASA NTRS1988

Experimental and theoretical analysis of the rate of solvent equilibration in the hanging drop method of protein crystal growth

The principles of the hanging-drop method of crystal growth are discussed, and the rate of water evaporation in a water droplet (containing protein, buffer, and a precipitating agent) suspended above a well containing a double concentration of precipitating agent is investigated theoretically. It is shown that, on earth, the rate of evaporation may be determined from diffusion theory and the colligative properties of solutions. The parameters affecting the rate of evaporation include the temperature, the vapor pressure of water, the ionization constant of the salt, the volume of the drop, the contact angle between the droplet and the coverslip, the number of moles of salt in the droplet, the number of moles of water and salt in the well, the molar volumes of water and salt, the distance from the droplet to the well, and the coefficient of diffusion of water vapor through air. To test the theoretical equations, hanging-drop experiments were conducted using various reagent concentrations in 25-microliter droplets and measuring the evaporation times at 4 C and 25 C. The results showed good agreement with the theory.

Fowlis, William W.↗
NASA NTRS1988

Cell separation by immunoaffinity partitioning with polyethylene glycol-modified Protein A in aqueous polymer two-phase systems

Previous work has shown that polyethylene glycol (PEG)-bound antibodies can be used as affinity ligands in PEG-dextran two-phase systems to provide selective partitioning of cells to the PEG-rich phase. In the present work it is shown that immunoaffinity partitioning can be simplified by use of PEG-modified Protein A which complexes with unmodified antibody and cells and shifts their partitioning into the PEG-rich phase, thus eliminating the need to prepare a PEG-modified antibody for each cell type. In addition, the paper provides a more rigorous test of the original technique with PEG-bound antibodies by showing that it is effective at shifting the partitioning of either cell type of a mixture of two cell populations.

Karr, Laurel J.↗
NASA NTRS1985

Regulation of protein degradation in muscle by calcium

Calcium-dependent regulation of intracellular protein degradation was studied in isolated rat skeletal muscles incubated in vitro in the presence of a large variety of agents known to affect calcium movement and distribution. The effect of different classes of protease inhibitors was tested to determine the responsible proteolytic systems involved in calcium-dependent degradation. The results suggest that nonlysosomal leupetin- and E-64-c-sensitive proteases are resposible for calcium-dependent proteolysis in muscle.

Zeman, Richard J.↗
NASA NTRS1989

Protein crystal growth in low gravity

The mechanisms involved in protein crystallization and those parameters which influence the growth process and crystalline perfection were studied. The analysis of the flows around growing crystals is detailed. The preliminary study of the growth of isocitrate lyase and the crystal morphologies found are discussed. Preliminary results of controlled nucleation studies are presented.

Feigelson, Robert S.↗
NASA NTRS1990

The Chemical Basis for the Origin of the Genetic Code and the Process of Protein Synthesis

A model for the origin of protein synthesis. The essential features of the model are that 5'-AMP and perhaps other monoribonucleotides can serve as catalysts for the selective synthesis of L-based peptides. A unique set of characteristics of 5'-AMP is responsible for the selective catalysts and these characteristics are described in detail. The model involves the formation of diesters as intermediates and selectivity for use of the L-isomer occurs principally at the step of forming the diester. However, in the formation of acetyl phenylalanine-AMP monoester there is a selectivity for esterification by the D-isomer. Data showing this selectivity is presented. This selectivity for D-isomer disappears after the first step. The identity was confirmed of all four of possible diesters of acetyl-D- and -L phenylaline with 5'-AMP by nuclear magnetic resonance (NMR). The data using flourescence and NMR show the Trp ring can associate with the adenine ring more strongly when the D-isomer is in the 2' position than it can when in the 3' position. These same data also suggest a molecular mechanisim for the faster esterificaton of 5'-AMP by acetyl-D-phenylaline. Some new data is also presented on the possible structure of the 2' isomer of acetyl-D-tryptophan-AMP monoester. The HPLC elution times of all four possible acetyl diphenylalanine esters of 5'-AMP were studied, these peptidyl esters will be products in the studies of peptide formation on the ribose of 5'-AMP. Other studies were on the rate of synthesis and the identity of the product when producing 3'Ac-Phe-2'tBOC-Phe-AMP diester. HPLC purification and identification of this product were accomplished.

Lacey, James C., Jr.↗
NASA NTRS1991

Compact Apparatus For Growth Of Protein Crystals

Compact apparatus proposed specifically for growth of protein crystals in microgravity also used in terrestrial laboratories to initiate and terminate growth at prescribed times automatically. Has few moving parts. Also contains no syringes difficult to clean, load, and unload and introduces contaminant silicon grease into crystallization solution. After growth of crystals terminated, specimens retrieved and transported simply.

Carter, Daniel C.↗
NASA NTRS1991

Metabolism of Nonessential N15-Labeled Amino Acids and the Measurement of Human Whole-Body Protein Synthesis Rates

Eight N-15 labeled nonessential amino acids plus (15)NH4Cl were administered over a 10 h period to four healthy adult males using a primed-constant dosage regimen. The amount of N-15 excreted in the urine and the urinary ammonia, hippuric acid, and plasma alanine N-15 enrichments were measured. There was a high degree of consistency across subjects in the ordering of the nine compounds based on the fraction of N-15 excreted (Kendall coefficient of concordance W = 0.83, P is less than 0.01). Protein synthesis rates were calculated from the urinary ammonia plateau enrichment and the cumulative excretion of N-15. Glycine was one of the few amino acids that gave similar values by both methods.

Stein, T. P.↗
NASA NTRS1991

Effect of denervation or unweighting on GLUT-4 protein in rat soleus muscle

The study is intended to test the hypothesis that the decreased capacity for glucose transport in the denervated rat soleus and the increased capacity for glucose transport in the unweighted rat soleus are related to changes in the expression of the regulatable glucose transporter protein in skeletal muscle (GLUT-4). Results obtained indicate that altered GLUT-4 expression may be a major contributor to the changes in insulin-stimulated glucose transport that are observed with denervation and unweighting. It is concluded that muscle activity is an important factor in the regulation of the GLUT-4 expression in skeletal muscle.

Henriksen, Erik J.↗
NASA NTRS1992

Protein crystal growth in low gravity

A study is presented of the crystallization of isocitrate lyase (ICL) and the influence of the lack of thermal solutal convection in microgravity on the morphology of ICL crystals is discussed. The latest results of studies with thermonucleation are presented. These include the nucleation of a protein with retrograde solubility and an unknown solubility curve. A new design for a more microgravity compatible thermonuclear is presented.

Feigelson, Robert S.↗