Search NASA⌕ Search

SEARCH · Search NASA

Results for “Isoelectric points”

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.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 records
All sourcesNASA NTRSDOE OSTI
NASA NTRS1983

Influence of deflocculant on the isoelectric point of refractory powders: Considerations on the action of deflocculant

Isoelectric point changes in suspensions of refractory materials vis-a-vis the role of deflocculants used in monolithic refractories were investigated by considering the mineral compositions and adsorbed ions in four kinds of clay. Three types of curves represented the relation between the isoelectric point and the deflocculant. The surface charge of clay particles in the suspensions became negative as a result of the deflocculant, since the isoelectric point of suspensions decreased as the deflocculant was added. The isoelectric point changes of calcined alumina were also compared with those of the clays, and a similar phenomenon was observed, except that the deflocculant dispersed the calcined alumina better than it did the clays. A simple model was used to analyze the results.

Naruse, Y.↗
NASA NTRS2018

Marginal protein stability drives subcellular proteome isoelectric point

There exists a positive correlation between the pH of subcellular compartments and the median isoelectric point (pI) for the associated proteomes. Proteins in the human lysosome—a highly acidic compartment in the cell—have a median pI of ∼6.5, whereas proteins in the more basic mitochondria have a median pI of ∼8.0. Proposed mechanisms reflect potential adaptations to pH. For example, enzyme active site general acid/base residue pKs are likely evolved to match environmental pH. However, such effects would be limited to a few residues on specific proteins, and might not affect the proteome at large. A protein model that considers residue burial upon folding recapitulates the correlation between proteome pI and environmental pH. This correlation can be fully described by a neutral evolution process; no functional selection is included in the model. Proteins in acidic environments incur a lower energetic penalty for burying acidic residues than basic residues, resulting in a net accumulation of acidic residues in the protein core. The inverse is true under alkaline conditions. The pI distributions of subcellular proteomes are likely not a direct result of functional adaptations to pH, but a molecular spandrel stemming from marginal stability.

Kaiser Loell↗
NASA NTRS1996

Characterization of the proteins comprising the integral matrix of Strongylocentrotus purpuratus embryonic spicules

In the present study, we enumerate and characterize the proteins that comprise the integral spicule matrix of the Strongylocentrotus purpuratus embryo. Two-dimensional gel electrophoresis of [35S]methionine radiolabeled spicule matrix proteins reveals that there are 12 strongly radiolabeled spicule matrix proteins and approximately three dozen less strongly radiolabeled spicule matrix proteins. The majority of the proteins have acidic isoelectric points; however, there are several spicule matrix proteins that have more alkaline isoelectric points. Western blotting analysis indicates that SM50 is the spicule matrix protein with the most alkaline isoelectric point. In addition, two distinct SM30 proteins are identified in embryonic spicules, and they have apparent molecular masses of approximately 43 and 46 kDa. Comparisons between embryonic spicule matrix proteins and adult spine integral matrix proteins suggest that the embryonic 43-kDa SM30 protein is an embryonic isoform of SM30. An adult 49-kDa spine matrix protein is also identified as a possible adult isoform of SM30. Analysis of the SM30 amino acid sequences indicates that a portion of SM30 proteins is very similar to the carbohydrate recognition domain of C-type lectin proteins.

Non-NASA Center↗
NASA NTRS1991

Purification and characterization of ornithine transcarbamylase from pea (Pisum sativum L.)

Pea (Pisum sativum) ornithine transcarbamylase (OTC) was purified to homogeneity from leaf homogenates in a single-step procedure, using delta-N-(phosphonacetyl)-L-ornithine-Sepharose 6B affinity chromatography. The 1581-fold purified OTC enzyme exhibited a specific activity of 139 micromoles citrulline per minute per milligram of protein at 37 degrees C, pH 8.5. Pea OTC represents approximately 0.05% of the total soluble protein in the leaf. The molecular weight of the native enzyme was approximately 108,200, as estimated by Sephacryl S-200 gel filtration chromatography. The purified protein ran as a single molecular weight band of 36,500 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These results suggest that the pea OTC is a trimer of identical subunits. The overall amino acid composition of pea OTC is similar to that found in other eukaryotic and prokaryotic OTCs, but the number of arginine residues is approximately twofold higher. The increased number of arginine residues probably accounts for the observed isoelectric point of 7.6 for the pea enzyme, which is considerably more basic than isoelectric point values that have been reported for other OTCs.

Non-NASA Center↗
NASA NTRS1990

Partial purification and characterization of a Ca(2+)-dependent protein kinase from the green alga, Dunaliella salina

A calcium-dependent protein kinase was partially purified and characterized from the green alga Dunaliella salina. The enzyme was activated at free Ca2+ concentrations above 10(-7) molar. and half-maximal activation was at about 3 x 10(-7) molar. The optimum pH for its Ca(2+)-dependent activity was 7.5. The addition of various phospholipids and diolein had no effects on enzyme activity and did not alter the sensitivity of the enzyme toward Ca2+. The enzyme was inhibited by calmodulin antagonists, N-(6-aminohexyl)-1-naphthalene sulfonamide and N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide in a dose-dependent manner while the protein kinase C inhibitor, sphingosine, had little effect on enzyme activity up to 800 micromolar. Immunoassay showed some calmodulin was present in the kinase preparations. However, it is unlikely the kinase was calmodulin regulated, since it still showed stimulation by Ca2+ in gel assays after being electrophoretically separated from calmodulin by two different methods. This gel method of detection of the enzyme indicated that a protein band with an apparent molecular weight of 40,000 showed protein kinase activity at each one of the several steps in the purification procedure. Gel assay analysis also showed that after native gel isoelectric focusing the partially purified kinase preparations had two bands with calcium-dependent activity, at isoelectric points 6.7 and 7.1. By molecular weight, by isoelectric point, and by a comparative immunoassay, the Dunaliella kinase appears to differ from at least some of the calcium-dependent, but calmodulin and phospholipid independent kinases described from higher plants.

Non-NASA Center↗
NASA NTRS1993

Characterization of auxin-binding proteins from zucchini plasma membrane

We have previously identified two auxin-binding polypeptides in plasma membrane (PM) preparations from zucchini (Cucurbita pepo L.) (Hicks et al. 1989, Proc. Natl. Acad. Sci. USA 86, 4948-4952). These polypeptides have molecular weights of 40 kDa and 42 kDa and label specifically with the photoaffinity auxin analog 5-N3-7-3H-IAA (azido-IAA). Azido-IAA permits both the covalent and radioactive tagging of auxin-binding proteins and has allowed us to characterize further the 40-kDa and 42-kDa polypeptides, including the nature of their attachment to the PM, their relationship to each other, and their potential function. The azido-IAA-labeled polypeptides remain in the pelleted membrane fraction following high-salt and detergent washes, which indicates a tight and possibly integral association with the PM. Two-dimensional electrophoresis of partially purified azido-IAA-labeled protein demonstrates that, in addition to the major isoforms of the 40-kDa and 42-kDa polypeptides, which possess isoelectric points (pIs) of 8.2 and 7.2, respectively, several less abundant isoforms that display unique pIs are apparent at both molecular masses. Tryptic and chymotryptic digestion of the auxin-binding proteins indicates that the 40-kDa and 42-kDa polypeptides are closely related or are modifications of the same polypeptide. Phase extraction with the nonionic detergent Triton X-114 results in partitioning of the azido-IAA-labeled polypeptides into the aqueous (hydrophilic) phase. This apparently paradoxical behavior is also exhibited by certain integral membrane proteins that aggregate to form channels. The results of gel filtration indicate that the auxin-binding proteins do indeed aggregate strongly and that the polypeptides associate to form a dimer or multimeric complex in vivo. These characteristics are consistent with the hypothesis that the 40-kDa and 42-kDa polypeptides are subunits of a multimeric integral membrane protein which has an auxin-binding site, and which may possess transporter or channel function.

NASA Program Space Biology↗
NASA NTRS1988

Ionic binding properties of carrier ampholytes

Radioactive ampholytes were synthesized with specific activity of 638 microCi/g. These were used in studies of ampholyte binding to target proteins under nonionic conditions. These radioactive ampholytes bound to target proteins but were dissociable in sodium chloride solutions with dissociation occurring in a concentration dependent way. The ampholytes could be dissociated from target molecules using excess unlabelled ampholytes synthesized in the laboratory as well as commercial ampholytes. Radioactive ampholytes were bound to target proteins with different isoelectric points, and the bound ampholytes were eluted and analyzed by recycling isoelectric focusing. The results showed that acidic proteins bound basic ampholytes and basic proteins bound acidic ampholytes.

Rodkey, L. Scott↗
NASA NTRS1992

Phosphorylation of the budgerigar fledgling disease virus major capsid protein VP1

The structural proteins of the budgerigar fledgling disease virus, the first known nonmammalian polyomavirus, were analyzed by isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The major capsid protein VP1 was found to be composed of at least five distinct species having isoelectric points ranging from pH 6.45 to 5.85. By analogy with the murine polyomavirus, these species apparently result from different modifications of an initial translation product. Primary chicken embryo cells were infected in the presence of 32Pi to determine whether the virus structural proteins were modified by phosphorylation. SDS-PAGE of the purified virus structural proteins demonstrated that VP1 (along with both minor capsid proteins) was phosphorylated. Two-dimensional analysis of the radiolabeled virus showed phosphorylation of only the two most acidic isoelectric species of VP1, indicating that this posttranslational modification contributes to VP1 species heterogeneity. Phosphoamino acid analysis of 32P-labeled VP1 revealed that phosphoserine is the only phosphoamino acid present in the VP1 protein.

Non-NASA Center↗
NASA NTRS1981

Mathematical modeling and computer simulation of isoelectric focusing with electrochemically defined ampholytes

A mathematical model of isoelectric focusing at the steady state has been developed for an M-component system of electrochemically defined ampholytes. The model is formulated from fundamental principles describing the components' chemical equilibria, mass transfer resulting from diffusion and electromigration, and electroneutrality. The model consists of ordinary differential equations coupled with a system of algebraic equations. The model is implemented on a digital computer using FORTRAN-based simulation software. Computer simulation data are presented for several two-component systems showing the effects of varying the isoelectric points and dissociation constants of the constituents.

Palusinski, O. A.↗
NASA NTRS1982

Isoelectric focusing in space

The potential of space electrophoresis is conditioned by the fact that all electrophoretic techniques require the suppression of gravity-caused convection. Isoelectric focusing (IEF) is a powerful variant of electrophoresis, in which amphoteric substances are separated in a pH gradient according to their isoelectric points. A new apparatus for large scale IEF, utilizing a recycling principle, has been developed. In the ground-based prototype, laminar flow is provided by a series of parallel filter elements. The operation of the apparatus is monitored by an automated array of pH and ultraviolet absorption sensors under control of a desk-top computer. The apparatus has proven to be useful for the purification of a variety of enzymes, snake venom proteins, peptide hormones, and other biologicals, including interferon produced by genetic engineering techniques. In planning for a possible space apparatus, a crucial question regarding electroosmosis needs to be addressed To solve this problem, simple focusing test modules are planned for inclusion in an early Shuttle flight.

Bier, M.↗
NASA NTRS1986

Utilization of non-conventional systems for conversion of biomass to food components: Recovery optimization and characterizations of algal proteins and lipids

Protein isolate obtained from green algae (Scenedesmus obliquus) cultivated under controlled conditions was characterized. Molecular weight determination of fractionated algal proteins using SDS-polyacrylamide gel electrophoresis revealed a wide spectrum of molecular weights ranging from 15,000 to 220,000. Isoelectric points of dissociated proteins were in the range of 3.95 to 6.20. Amino acid composition of protein isolate compared favorably with FAO standards. High content of essential amino acids leucine, valine, phenylalanine and lysine makes algal protein isolate a high quality component of closed environment life support system (CELSS) diets. To optimize the removal of algal lipids and pigments supercritical carbon dioxide extraction (with and without ethanol as a co-solvent) was used. Addition of ethanol to supercritical CO2 resulted in more efficient removal of algal lipids and produced protein isolate with a good yield and protein recovery. The protein isolate extracted by the above mixture had an improved water solubility.

Karel, M.↗
NASA NTRS1987

Non-conventional approaches to food processing in CELSS, 1. Algal proteins: Characterization and process optimization

Protein isolate obtained from green algae cultivated under controlled conditions was characterized. Molecular weight determination of fractionated algal proteins using SDS-polyacrylamide gel electrophoresis revealed a wide spectrum of molecular weights ranging from 15,000 to 220,000. Isoelectric points of dissociated proteins were in the range of 3.95 to 6.20. Amino acid composition of protein isolate compared favorably with FAO standards. High content of essential amino acids leucine, valine, phenylalanine and lysine make algal protein isolate a high quality component of closed ecological life support system diets. To optimize the removal of algal lipids and pigments supercritical carbon dioxide extraction (with and without ethanol as a co-solvent) was used. Addition of ethanol to supercritical carbon dioxide resulted in more efficient removal of algal lipids and produced protein isolate with a good yield and protein recovery. The protein isolate extracted by the above mixture had an improved water solubility.

Nakhost, Z.↗
NASA NTRS1987

Non-conventional approaches to food processing in CELSS. I - Algal proteins: Characterization and process optimization

Protein isolate obtained from green algae (Scenedesmus obliquus) cultivated under controlled conditions was characterized. Molecular weight determination of fractionated algal proteins using SDS-polyacrylamide gel electrophoresis revealed a wide spectrum of molecular weights ranging from 15,000 to 220,000. Isoelectric points of dissociated proteins were in the range of 3.95 to 6.20. Amino acid composition of protein isolate compared favorably with FAO standards. High content of essential amino acids leucine, valine, phenylalanine and lysine makes algal protein isolate a high quality component of CELSS diets. To optimize the removal of algal lipids and pigments supercritical carbon dioxide extraction (with and without ethanol as a co-solvent) was used. Addition of ethanol to supercritical CO2 resulted in more efficient removal of algal lipids and produced protein isolate with a good yield and protein recovery. The protein isolate extracted by the above mixture had an improved water solubility.

Nakhost, Z.↗
NASA NTRS1988

Comparison of bioseparation methods for microgravity experiments

The efficiency of the 1-g version of the continuous-flow electrophoresis (CFE) system flown on Space Shuttle missions is compared with the efficiency of a commercial CFE for separating living cells (human kidney, liver, and pituitary-gland cells and T-lymphocytes). In addition, the CFE system and a reciprocal isoelectric focusing (RIEF) system are compared with respect to protein pyrification efficiency. Correlations were made among electrophoretic mobilities (EPMs), secretory functions of cells, and input sample concentrations. A significant reduction in mean and range EPM was observed when input sample concentrations exceeded a low threshohold. This effect was not observed in microgravity experiments conducted at sample concentrations three times greater than the threshold for the controls. Comparison of CFE and RIEF methods showed that there are apparent advantages for each method depending on the product. For example, RIEF purification of urokinase removed more protein impurities, but focused the enzyme at a pH different than the enzyme's known isoelectric point.

Morrison, Dennis R.↗
NASA NTRS1989

A comparison of an ATPase from the archaebacterium Halobacterium saccharovorum with the F1 moiety from the Escherichia coli ATP Synthase

A purified ATPase associated with membranes from Halobacterium saccharovorum was compared with the F sub 1 moiety from the Escherichia coli ATP Synthase. The halobacterial enzyme was composed of two major (I and II) and two minor subunits (III and IV), whose molecular masses were 87 kDa, 60 kDa, 29 kDa, and 20 kDa, respectively. The isoelectric points of these subunits ranged from 4.1 to 4.8, which in the case of the subunits I and II was consistent with the presence of an excess of acidic amino acids (20 to 22 Mol percent). Peptide mapping of sodium dodecylsulfate-denatured subunits I and II showed no relationship between the primary structures of the individual halobacterial subunits or similarities to the subunits of the F sub 1 ATPase (EC 3.6.1.34) from E. coli. Trypsin inactivation of the halobacterial ATPase was accompanied by the partial degradation of the major subunits. This observation, taken in conjunction with molecular masses of the subunits and the native enzyme, was consistent with the previously proposed stoichiometry of 2:2:1:1. These results suggest that H. saccharovorum, and possibly, Halobacteria in general, possess an ATPase which is unlike the ubiquitous F sub o F sub 1 - ATP Synthase.

Stan-Lotter, Helga↗
NASA NTRS1998

The Development of an Edible Peanut Protein Film

The peanut is one of the crops chosen for use in NASA's Advanced Life Support Program (ALS). The peanut is a source of both oil and protein. After oil is extracted from the peanut, a protein rich flour remains. An edible peanut protein film is one use for this flour. Two types of film are developed for this study, one set of film contains 10% fat while the other set contains no fat. For film without fat the defatting of the peanut by the Soxhlet method is the first step in the manufacturing process of the film. Secondly, the protein is precipitated at its isoelectric point (pH 4.5) and centrifuged to separate the protein from the non-protein. After freeze-drying the protein it is milled in a ceramic ball mill to decrease particle size and sifted through a series of sieves to determine particle size distribution. Those particles retained on the 100 mesh sieves are utilized for film formation. Larger particles are re-ground and sifted. Five grams of protein is mixed with 50 mL of distilled water, 70 mL of 80% ethanol, 15 mL of 6N ammonium hydroxide and a plasticizer. This mixture is heated for 30 minutes until the temperature reaches 70 C. The mixture is then poured onto a level Teflon coated glass surface. After allowing the film to form overnight under a ventilation hood, it is manually removed from the plate. The processes and methods adopted have created flexible films of uniform thickness that are free of air bubbles. Thickness of films made from defatted peanut protein and partially defatted peanut protein were 0.10 Lm and 0.13 Lm respectively. Films with natural peanut fat are approximately three times as flexible and almost four times as strong as the films made without fat. Further research will be performed to evaluate its mechanical properties. This paper will greatly contribute to food preservation and waste management. Potential applications of this film are edible/biodegradable containers, wrapping for food preservation (against water, oxygen and oil), storage packets for seasonings or other ingredients and encapsulation for pharmaceutical use.

Patrick, N.↗
NASA NTRS1990

Partial purification and characterization of indol-3-ylacetylglucose:myo-inositol indol-3-ylacetyltransferase (indoleacetic acid-inositol synthase)

A procedure is described for the purification of the enzyme indol-3-ylacetylglucose:myo-inositol indol-3-ylacetyltransferase (IAA-myo-inositol synthase). This enzyme catalyzes the transfer of indol-3-ylacetate from 1-0-indol-3-ylacetyl-beta-d-glucose to myo-inositol to form indol-3-ylacetyl-myo-inositol and glucose. A hexokinase or glucose oxidase based assay system is described. The enzyme has been purified approximately 16,000-fold, has an isoelectric point of pH 6.1 and yields three catalytically inactive bands upon acrylamide gel electrophoresis of the native protein. The enzyme shows maximum transferase activity with myo-inositol but shows some transferase activity with scyllo-inositol and myo-inosose-2. No transfer of IAA occurs with myo-inositol-d-galactopyranose, cyclohexanol, mannitol, or glycerol as acyl acceptor. The affinity of the enzyme for 1-0-indol-3-ylacetyl-beta-d-glucose is, Km = 30 micromolar, and for myo-inositol is, Km = 4 millimolar. The enzyme does not catalyze the exchange incorporation of glucose into IAA-glucose indicating the reaction mechanism involves binding of IAA glucose to the enzyme with subsequent hydrolytic cleavage of the acyl moiety by the hydroxyl of myo-inositol to form IAA myo-inositol ester.

NASA Program Space Biology↗
NASA NTRS1988

Enzymic synthesis of indole-3-acetyl-1-O-beta-d-glucose. I. Partial purification and characterization of the enzyme from Zea mays

The first enzyme-catalyzed reaction leading from indole-3-acetic acid (IAA) to the myo-inositol esters of IAA is the synthesis of indole-3-acetyl-1-O-beta-D-glucose from uridine-5'-diphosphoglucose (UDPG) and IAA. The reaction is catalyzed by the enzyme, UDPG-indol-3-ylacetyl glucosyl transferase (IAA-glucose-synthase). This work reports methods for the assay of the enzyme and for the extraction and partial purification of the enzyme from kernels of Zea mays sweet corn. The enzyme has an apparent molecular weight of 46,500 an isoelectric point of 5.5, and its pH optimum lies between 7.3 and 7.6. The enzyme is stable to storage at zero degrees but loses activity during column chromatographic procedures which can be restored only fractionally by addition of column eluates. The data suggest either multiple unknown cofactors or conformational changes leading to activity loss.

NASA Discipline Number 40-10↗