Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two acid proteases were isolated from the soluble extracts of adult Dirofilaria immitis, the filarial heartworm of canines. Activity of these proteases was detected using 3H-labeled bovine alpha-casein as substrate, and they were designated Fp-I and Fp-II in order of their elution from a CM-cellulose column. The molecular weight of partially purified Fp-I was approximately 170000, and it was active between pH 4.6-5.8. The activity of Fp-I doubled in the presence of various sulfhydryl reagents at 5 mM, and it was inhibited 50-60% by the sulfhydryl inhibitors p-hydroxymercuribenzoate and iodoacetate at 1 mM, the heavy metal chelating agent o-phenanthroline at 1 mM and the peptide aldehyde protease inhibitors pepstatin (10 microM), leupeptin, antipain and chymostatin (50 microM). The molecular weight of the more extensively purified Fp-II is approximately 48000. This protease was active between pH 2.6-3.4 and was highly sensitive to inhibition by pepstatin (80% inhibition at 10 nM). Fp-II was not significantly affected by sulfhydryl reagents, sulfhydryl inhibitors, metal chelating agents or peptide aldehyde protease inhibitors other than pepstatin. These properties of dirofilarial Fp-II resemble those of mammalian cathepsin D.
Mol Biochem Parasitol 1983 Sep
PMID:Isolation, partial purification and some properties of two acid proteases from adult Dirofilaria immitis. 636 44

A new method of preparing biological samples for electron microscopy has been used to re-examine the structure of actin filaments, actin filaments decorated by myosin subfragment-1 (S1), and insect flight muscles. Samples were quick-frozen by contact with a block of copper cooled to approximately 4 K; then were freeze-fractured, deep-etched, rotary-replicated with platinum, and viewed in a transmission electron microscope. By this approach, actin filaments display prominent transverse bands whose repeat (approximately 5.5 nm) and pitch (approximately 15 to 20 degrees) fit with the expected left-handed "genetic" helix. Freeze-etched actin filaments do not, however, display the usual two-start helix as prominently as is seen after negative staining, and they also appear substantially thicker than after negative staining (9 to 10 nm versus 8 nm). The latter two-start helix appears very clearly after S1 decoration. Nevertheless, freeze-etched acto-S1 does not display the "arrowheads" that are seen after negative staining. Instead it displays the outer envelope of the helically deployed S1, and as would be expected from current models derived from optical reconstruction of negatively stained samples, this surface view looks only slightly polarized. Finally, the quick-freeze, deep-etch approach provides particularly distinct images of the crossbridges in insect flight muscles. These are plentiful and regularly arranged in rigor muscles, but rare in muscles relaxed with ATP before freezing. In rigor muscles fixed with aldehydes, these crossbridges assume a broad distribution of inclination, ranging from 45 degrees to 90 degrees with a mean of approximately 80 degrees, which is less tilt than has been seen before in thin-sectioned muscles. However, when aldehyde fixation is followed by exposure to tannic acid with or without uranyl acetate block-staining, crossbridges assume a more acute angle with respect to the fiber axis, centering around 45 degrees. This is associated with a commensurate reduction in interfilament spacing within the muscle fibers, such that tilted crossbridges are not any longer than untilted ones (both measuring approximately 15 nm). At the opposite extreme, crossbridges often become stretched in unfixed muscles, owing to an unnatural increase in interfilament spacing that occurs during sample preparation; in such regions, crossbridges display narrow "stalks", which invariably emerge from the thick filaments at close to 90 degrees. We conclude that crossbridge shape and orientation is strongly affected by different methods of sample preparation, and this will make it difficult to visualize natural crossbridge movements by electron microscopy.
J Mol Biol 1983 Sep 05
PMID:Actin-myosin interactions visualized by the quick-freeze, deep-etch replica technique. 662 Mar 83

To provide for bioluminescence measurements of the enzymatic activities of dehydrogenases, disturbing contaminants were removed from a bacterial luciferase extract by chromatography, using Blue Sepharose CL-6B, a cross-linked agarose to which Cibacrone Blue F3G-A is covalently attached. This compound has a strong affinity to the dinucleotide fold, which is a region in enzymes binding NAD(H) or NADP(H). In contrast to the absorbed dehydrogenases, both luciferase and oxidoreductase were easily eluted and appeared close to the main bulk of UV-absorbing but analytically less important material. A rapid recording of the elution of luciferase was accomplished with a new electrochemical bioluminescence assay. Due to this and the early elution of the desired material, it could be chromatographed, recognized and collected in less than two hours. Thereby the light-yielding capacity of the sensitive material was well preserved. For bioluminescence assay solutions composed of pooled oxidoreductase-luciferase fractions, FMN and a long chain aldehyde were prepared and supplemented with NAD+ and either lactate, malate or 3-hydroxybutyrate. The analyses were carried out in a single step performance by adding the enzyme sample to the luciferase solution. Minute amounts of lactate dehydrogenase, malate dehydrogenase and 3-hydroxybutyrate dehydrogenase yielded a linear light response permitting assay in the lower part of the femtomole region. In case a dehydrogenase does not occur as a contaminant of a commercial luciferase preparation, purification with Cibacrone Blue can be omitted as demonstrated for glucose-6-phosphate dehydrogenase.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Cell Biochem 1983
PMID:Single-step bioluminescence analyses of enzymes, using Cibacrone Blue chromatography for removal of interfering dehydrogenases. 663 14

A proteolytic enzyme of the liver fluke Fasciola sp. was purified as described previously by ammonium sulfate fractionation, gel filtration on Sephadex G-200 column and L-phenylalanine-agarose chromatography. Leupeptin, a peptide aldehyde of microbial origin, competitively inhibited the enzyme activity with respect to the substrate alpha-N-benzoyl-L-argininamide; the apparent Ki value for leupeptin is 45 000-fold less than the apparent Km for the substrate. Incubation of the enzyme with leupeptin resulted in time-dependent inactivation of the globinolytic activity, with an inactivation constant (Kinact) of 0.4 microM giving the half-maximum inactivation velocity, and with a minimum inactivation half-time (T) of 2.7 min at infinite concentration of this compound. The inactivated enzyme was not reactivated by extensive dialysis. These results imply that leupeptin yields an affinity labelling of an active site of the enzyme. The activity of the Fasciola proteolytic enzyme was also inactivated by other peptide aldehydes and alkylating agents and inactivation constants observed were 0.5 microM for chymostatin, 13 microM for antipain, 2 microM for p-toluenesulfonyl-L-lysine chloromethyl ketone, 140 microM for p-toluenesulfonyl-L-phenylalanine chloromethyl ketone and 40 microM for iodoacetate under the conditions used.
Mol Biochem Parasitol 1983 Jun
PMID:Mechanism of inactivation of a Fasciola proteolytic enzyme by peptide aldehydes and alkylating agents. 687 85

One hypothesis of the origin of pre-modern forms of life is that the original replicating molecules were specific polypeptides which acted as templates for the assembly of poly-Schiff bases complementary to the template, and that these polymers were then oxidized to peptide linkages, probably by photo-produced oxidants. A double cycle of such anti-parallel complementary replication would yield the original peptide polymer. If this model were valid, the Schiff base between an N-acyl alpha amino aldehyde and an amino acid should yield a dipeptide in aqueous solution in the presence of an appropriate oxidant. In the present study it is shown that the substituted dipeptide, N-acetyl-tyrosyl-tyrosine, is produced in high yield in aqueous solution at pH 9 through the action of H2O2 on the Schiff-base complex between N-acetyl-tyrosinal and tyrosine and that a great variety of N-acyl amino acids are formed from amino acids and aliphatic aldehydes under similar conditions.
J Mol Evol 1982
PMID:Oxidative peptide (and amide) formation from Schiff base complexes. 716 6

Oxygen-free aqueous solutions of CH3CN (0.1 M, pH 6) were exposed to gamma rays from a 60Co source, the mixture of nonvolatile radiolytic products was fractionated and the fractions were analysed. Succinic, maleic, fumaric, malonic and pyruvic acids were identified. Glycol aldehyde, glucose and probably ribose were observed in the hydrolysate of fractionated material. It has been suggested that an oligomer is formed which has a fragment with the polyhydroxy structure and on hydrolysis releases the carbohydrates. Radiolytic products which release amino acids on hydrolysis were found in several fractions. The amino acid contents of the hydrolysates were up to about 2.8% of the fraction mass. The presence of several protein and nonprotein amino acids suggests that their origin should be in a peptidic structure, which is probably a fragment of an oligomer radiolytically produced. A direct analysis of the irradiated solution shows the presence of acetaldehyde, propionaldehyde, glyoxal and of biacetyl. Experimental findings are discussed and a free-radical mechanism is proposed to account for the chemical changes observed.
J Mol Evol 1980 Jul
PMID:The radiolysis of aqueous acetonitrile: compounds of interest to chemical evolution studies. 740 Nov 81

The authors have recently described the development of a carboxymethyl dextran-based sensor surface for biospecific interaction analysis by surface plasmon resonance. Ligands are immobilized via primary amine groups after activation of the carboxymethyl groups on the sensor surface with a mixture of N-hydroxysuccinimide and N-ethyl-N'-(dimethylaminopropyl) carbodiimide. Methods have now been developed for efficient immobilization via thiol/disulfide exchange, aldehyde coupling and biotin-avidin coupling. The specific activity of monoclonal antibodies immobilized by the four different methods was investigated by altering the immobilization conditions, e.g., activation time, protein concentration, ionic strength and the degree of modification, etc. Investigations have also been made concerning possible differences in the specific activity for antibodies immobilized using optimized conditions with respect to the four different chemistries. These studies show that, with the flexible carboxymethyl dextran matrix used here, the immobilization methods give rise to only minor differences in specific activity. Thus, with this solid support, a 'site directed' immobilization strategy for monoclonal antibodies has no advantage. In general the specific activity for optimized systems was approximately 75% for the binding of beta 2 mu-globulin to an immobilized monoclonal antibody directed against beta 2 mu-globulin. Reduced specific activities of immobilized antibodies induced by variation of the coupling conditions could be attributed to the deterioration of the active site of the antibody.
J Mol Recognit
PMID:Comparison of methods for immobilization to carboxymethyl dextran sensor surfaces by analysis of the specific activity of monoclonal antibodies. 754 Dec 26

Non-phosphorylating glyceraldehyde 3-phosphate dehydrogenase (GAPDH, NADP-specific, EC 1.2.1.9) operates in the cytosol of autotrophic eukaryotes where it generates NADPH for biosynthetic processes from photosynthetic glyceraldehyde 3-phosphate exported from the chloroplast by the phosphate translocator. Here we report the first cloning and characterization of cDNAs encoding complete polypeptide chains of nonphosphorylating GAPDH from pea and maize by using oligonucleotide probes derived from amino acid sequences determined for the purified enzyme. Unexpectedly, nonphosphorylating GAPDH cannot be aligned with the well-known sequences of phosphorylating GAPDH, but shares about 30% amino acid identity with various specialized and non-specialized aldehyde dehydrogenases (ALDHs) of eubacteria and eukaryotes. A phylogenetic analysis of this ALDH superfamily reveals a complex evolutionary pattern with numerous major branches carrying genes from eubacteria, eukaryotes, or both, encoding enzymes that are specific or non-specific for particular aldehyde substrates. This topology suggests a concomitant emergence of multiple substrate specificities from non-specialized ALDH during an early evolutionary phase of intense metabolic diversification. Although unrelated at the sequence level, non-phosphorylating aldehyde dehydrogenases and phosphorylating GAPDH resemble one another with respect to catalytic hydride transfer and covalent thiol ester formation. Whether or not this reflects an ancestral relationship can only be decided when crystallographic data for ALDH enzymes have become available.
J Mol Biol 1994 Mar 18
PMID:Non-phosphorylating GAPDH of higher plants is a member of the aldehyde dehydrogenase superfamily with no sequence homology to phosphorylating GAPDH. 754 14

In the present investigation we have shown that PBLs taken from HIV-positive patients suppressed T-cell proliferation when cultured for at least 12 days. When infected In vitro with HIV-1, these cells become suppressive after 6 days. PBLs collected from seronegative individuals turned out to be non-suppressive even after prolonged culturing. However, when infected in vitro with HIV they were found to be cytostatic also after 6 days. This anti-immune activity, related to HIV infection, is mediated, at least in part, by alpha IFN, since about 50% of this HIV associated activity could be quenched by alpha IFN antibodies. We also showed that this T-cell cytostasis is associated with CD8+ cells. In view of a possible use of suppressive cells for T-cell vaccination, we verified that such cellular functions were abolished by aldehyde treatment.
Cell Mol Biol (Noisy-le-grand) 1995 May
PMID:Contribution of alpha interferon (alpha IFN) to HIV-induced immunosuppression. 758 Aug 38

Ovarian produced ecdysteroids regulate sex pheromone production in the female housefly, inducing the synthesis of (Z)-9-tricosene (Z9-23:Hy), cis-9,10-epoxytricosane, (Z)-14-tricosen-10-one and methylalkanes. Experiments were performed to gain a detailed understanding of the processes affected by 20-hydroxyecdysone (20-HE) that result in sex pheromone production as the female becomes reproductively mature. A novel microsomal fatty acid synthetase (FAS) is present in the epidermal tissue and plays a role in producing the methyl-branched fatty acid precursors to the methylalkanes. This FAS is released from the microsomes in the presence of 3 M KCl. A major enzyme activity influenced by 20-HE is the fatty acyl-CoA elongation system. A shift in the chain length specificity of the products of the elongation system causes the change in the chain lengths of the alkenes produced to switch from C27 and longer in the previtellogenic female to C23 in the mature female. Data is presented indicating that it is the condensation activity of the elongation system that is affected. Z9-23:Hy arises from a 24 carbon acyl group which is reduced to an aldehyde, and then converted to the hydrocarbon. Data is presented demonstrating that it is the fatty acyl-CoA derivative and not the free fatty acid that is the substrate. There does not appear to be a chain length specificity which regulates the conversion of fatty acyl-CoAs to hydrocarbons as both 24 and 28 carbon fatty acyl-CoAs are converted to hydrocarbon by both males and females of all ages.
Insect Biochem Mol Biol 1995 Jun
PMID:Regulation of enzymatic activity involved in sex pheromone production in the housefly, Musca domestica. 762 7


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