UMLS:C0027960 - FACTA Search

Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Target Concepts:

Query: UMLS:C0027960 (mole)
21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This study characterizes the structural and functional significance of sulfhydryl residues in human plasma heparin cofactor II (HCII). For quantification of sulfhydryl groups, the extinction coefficient of HCII was redetermined and found to be 0.593 ml mg-1 cm-1 using second-derivative spectroscopy and multicomponent analysis assuming 4, 10, and 2 residues of tryptophan, tyrosine, and tyrosine-O-sulfate per mole of protein, respectively. The results show that tyrosine-O-sulfate residues in HCII and in cholecystokinin peptide fragments (as model compounds) do not significantly contribute to the absorbance spectrum from 280 to 300 nm. A total of three sulfhydryl groups per mole of HCII was detected by Ellman's reagent titration, with or without treatment with dithioerythritol, indicating the absence of intramolecular disulfide bonds. Incubation of HCII with 0.1-10 mM dithioerythritol did not diminish its heparin-enhanced thrombin inhibition activity. Treatment with various sulfhydryl-specific reagents, including p-mercuribenzoate, HgCl2, and N-substituted maleimide derivatives, inactivated HCII. Titration with Ellman's reagent after these reactions identified the modification site as a cysteinyl residue(s). However, complete methanethio derivatization of the sulfhydryl groups of HCII using methyl methanethiosulfonate did not alter heparin-catalyzed thrombin inhibition. These results indicate that the sulfhydryl groups of HCII are not essential for thrombin inhibition. HCII differs from antithrombin III, which contains an essential disulfide bond for heparin-dependent thrombin inhibition (Longas, M. O., et al. (1980) J. Biol. Chem. 255, 3436). Furthermore, within the "serpin" (serine proteinase inhibitor) superfamily, HCII resembles chicken ovalbumin in occurrence of sulfhydryl residues and reactivity with various sulfhydryl group-directed compounds.
...
PMID:Structure-function relationships in heparin cofactor II: spectral analysis of aromatic residues and absence of a role for sulfhydryl groups in thrombin inhibition. 342 30

Rabbit skeletal muscle calsequestrin was fragmented by using trypsin in the presence and absence of calcium. Calcium ion was found to protect calsequestrin from proteolysis, and the peptides produced in the presence of calcium were stable to further digestion. Peptides produced in the presence or absence of calcium had a decreased helical content but maintained their ability to bind calcium. The amino acid sequence of a 59-residue carboxyl-terminal tryptic peptide was determined by automated Edman degradation and carboxypeptidase Y digestion of carboxyl-terminal tryptic, chymotryptic, and cyanogen bromide peptides. This peptide is highly acidic (Asp + Glu = 42%, Lys + Arg = 0), and it bound a total of 15 calcium ions per mole of peptide (Kd = 8.5 mM). The intrinsic tryptophan fluorescence of the peptide was enhanced by 10% upon binding Ca2+ with the dissociation constant of 1 mM. Analyses of the circular dichroism spectra of the peptide showed that it was primarily in a random-coil conformation with little helical (2%) and moderate beta-structure (25%) regardless of the calcium concentration. This peptide also bound 7 mol of terbium per mole of peptide with high affinity (Kd = 7.5 microM).
...
PMID:Fragmentation of rabbit skeletal muscle calsequestrin: spectral and ion binding properties of the carboxyl-terminal region. 342 87

Two naturally occurring non-enzymic glucosylceramide activator proteins (A1a and A1b activator) shown previously to be immunochemically not detectable in a new variant of human Gaucher disease (glucosylceramide lipidosis) without glucosylceramidase deficiency, were characterized by amino-acid sequence and carbohydrate content. The complete amino-acid sequence of the A1a activator was determined. The protein consists of 80 amino-acid residues including three disulfide bridges lacking arginine and tryptophan. The molecular mass is 8.95 kDa. About 20% of the polypeptide chain are shorter by two amino-acid residues at the N-terminal end. The A1b activator was characterized by the amino-acid compositions of all tryptic peptides and of the entire protein; sequencing was performed of the regions 1-34 and 42-56. Identical results were obtained for the polypeptide chains of both A1 activators. This suggests that they do not differ in their primary structures which is in agreement with the immunochemical results. The difference between A1a and A1b activator is due to the carbohydrate part. The total amount of 49% carbohydrate in A1a and 76.7% in A1b consists mainly of hexoses. Both chains contain two moles of N-acetylglucosamine per mole protein bound to asparagine in position 22. A comparison of the primary structure of the A1 activator with the sulfatide activator sequence revealed an interesting similarity, especially of the cysteine residues and the carbohydrate-binding asparagine. Sequence homology was also found between a part of the A1 activator sequence and the hemagglutinin neuraminidase of influenza virus as well as to a hypothetical glycoprotein of the Epstein-Barr virus. The comparison with human lysosomal glucosylcerebrosidase showed no sequence similarity.
...
PMID:Complete amino-acid sequence and carbohydrate content of the naturally occurring glucosylceramide activator protein (A1 activator) absent from a new human Gaucher disease variant. 344

D-beta-Hydroxybutyrate dehydrogenase is a lipid-requiring enzyme with absolute specificity for phosphatidylcholine (PC). The enzyme devoid of lipid, the apodehydrogenase, inserts spontaneously into phospholipid vesicles where it exists as a tetramer. We now find the lipid activation to be limited by the mole fraction of PC in the total phospholipid. These studies suggest that the concentration of the enzyme-PC complex, which is essential for enzymic activity, becomes diffusion limited at lower PC concentration. The lipid activation and the tryptophan fluorescence of purified D-beta-hydroxybutyrate dehydrogenase were studied in the presence of a constant "bilayer background" of approximately 100 nonactivating phospholipid molecules/enzyme monomer. Activation by PC was half-maximal at 20 PC molecules/enzyme monomer. This value was doubled when the amount of "background" phospholipid was doubled. Activation proceeded with positive cooperativity having a Hill coefficient of approximately 2.4. These data indicate interactions between at least three PC-binding sites. The quenching of tryptophan fluorescence by the phospholipid activator, 1-palmitoyl-2-(1-pyrenyl)-decanoyl-PC (2-pyrenyl-PC), gives a saturation curve with half-maximal quenching of 6 quencher molecules/enzyme monomer. This value is equivalent to an apparent phospholipid-protein dissociation constant in the two-dimensional membrane and corresponds to approximately 6 mol % of total phospholipid. In distinct contrast to the phospholipid activation curve, the fluorescence quenching saturation curve was hyperbolic and there was no specificity for PC. The fluorescence quenching by 2-pyrenyl-PC could be diminished by using a several-fold excess of PC or other phospholipids so as to reduce the mole fraction of quencher in the bilayer. It would appear that formation of enzyme-PC complex is a dynamic process consisting of at least two discernible steps: 1) a primary interaction, as measured by tryptophan quenching, which is hyperbolic and not specific for lecithin. This interaction is independent from and precedes 2) phospholipid activation of D-beta-hydroxybutyrate dehydrogenase, which is cooperative in nature and specific for lecithin.
...
PMID:Site-site interaction in the phospholipid activation of D-beta-hydroxybutyrate dehydrogenase. 350 87

By gel filtration and titration on DEAE-cellulose filters we show that Escherichia coli tryptophanyl-tRNA synthetase forms tryptophanyl adenylate as an initial reaction product when the enzyme is mixed with ATP-Mg and tryptophan. This reaction precedes the synthesis of the tryptophanyl-ATP ester known to be formed by this enzyme. The stoichiometry of tryptophanyl adenylate synthesis is 2 mol per mole of dimeric enzyme. When this reaction is studied either by the stopped-flow method, by the fluorescence changes of the enzyme, or by radioactive ATP depletion, three successive chemical processes are identified. The first two processes correspond to the synthesis of the two adenylates, at very different rates. The rate constants of tryptophanyl adenylate synthesis are respectively 146 +/- 17 s-1 and 3.3 +/- 0.9 s-1. The third process is the synthesis of tryptophanyl-ATP, the rate constant of which is 0.025 s-1. The Michaelis constants for ATP and for tryptophan in the activation reaction are respectively 179 +/- 35 microM and 23.9 +/- 7.9 microM, for the fast site, and 116 +/- 45 microM and 3.7 +/- 2.2 microM, for the slow site. No synergy between ATP and tryptophan can be evidenced. The data are interpreted as showing positive cooperativity between the subunits associated with conformational changes evidenced by fluorometric methods. The pyrophosphorolysis of tryptophanyl adenylate presents a Michaelian behavior for both sites, and the rate constant of the reverse reaction is 360 +/- 10 s-1 with a binding constant of 196 +/- 12 microM for inorganic pyrophosphate (PPi).(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Tryptophanyl adenylate formation by tryptophanyl-tRNA synthetase from Escherichia coli. 351 15

The Salmonella typhimurium periplasmic histidine-binding J-protein is one of four proteins encoded by the histidine transport operon. Mutant J-protein hisJ5625 binds L-histidine, but does not transport it. The tertiary structure and conformational dynamics of native and mutant J-protein have been compared using steady state fluorescence, fluorescence polarization, and fluorescence energy transfer measurements. The two proteins have different three-dimensional structures and exhibit different responses to histidine binding. Ligand-induced conformational changes were demonstrated in both J-proteins using fluorescence energy transfer (distant reporter method) between the single tryptophan residue per mole of protein and a fluorescein-labeled methionine residue. However, the conformational change of the mutant protein is qualitatively and quantitatively different from that of the wild-type protein. Moreover, the microenvironment of the tryptophan and its distance from the labeled methionine (44A for the wild type, 60A for the mutant J-protein) are different in the two proteins. In conclusion, these results indicate that the specific conformational change induced in the wild type J-protein is a necessary requirement for the transport of L-histidine.
...
PMID:Conformational dynamics of two histidine-binding proteins of Salmonella typhimurium. 352 54

Human plasma glutathione peroxidase (GPx) was purified to homogeneity by ammonium sulfate fractionation, gel filtration on Sephadex G-150, chromatography on DEAE Sephacel, chromatofocusing with polybuffer, and gel filtration with Sephadex G-75. This isolation resulted in about 5,400-fold purification of the enzyme with a 32% yield in enzyme activity. The final preparation had a specific activity of about 28 units (mmoles NADPH oxidized) per milligram of protein. Determination of selenium on the purified enzyme revealed a content of 3.8 g atoms per mole GPx. Gel electrophoresis using SDS with standard proteins revealed a molecular weight of about 23,000 for the subunits, which would indicate a molecular weight of about 92,000 for the native enzyme. Amino acid analyses of the purified GPx indicated aspartate, glutamate, proline, glycine, alanine, and leucine as the predominant amino acids and cysteine, methionine, tryptophan, and histidine as the minor amino acids.
...
PMID:Properties of glutathione peroxidase isolated from human plasma. 366 26

Apparent kinetic constants (Km and Vmax values) were determined for human liver acyl-CoA: glycine acyltransferase (glycine-N-acylase) towards isobutyryl-CoA, 2-methyl butyryl-CoA, isovaleryl-CoA, butyryl-CoA, hexanoyl-CoA, octanoyl-CoA, and decanoyl-CoA. These acyl-CoA esters were selected because of their relevance to the human diseases with cellular accumulation of these esters, i.e., especially to metabolic defects in the acyl-CoA dehydrogenation steps of the branched-chain amino acids, lysine, 5-hydroxy lysine, tryptophan, and fatty acid oxidation pathways. With the acyl-CoA ester as the fixed substrate, the Km value for glycine ranged from 0.5 to 2.9 mole/liter, and with glycine as fixed substrate, the Km values for the acyl-CoA esters varied from 0.3 to 5.6 mmole/liter. It is concluded that the substrate concentration is decisive for the glycine conjugate formation and that the occurrence in urine of acylglycines reflects an intramitochondrial accumulation of the corresponding acyl-CoA ester.
...
PMID:Acyl-CoA: glycine N-acyltransferase: in vitro studies on the glycine conjugation of straight- and branched-chained acyl-CoA esters in human liver. 370 52

When tryptophanyl-tRNA synthetase from Escherichia coli is allowed to react with L-tryptophan and ATP-Mg in the presence of inorganic pyrophosphatase, the fluorescence change of the reaction mixture reveals three or four sequential processes, depending on the buffer used. Quenched-flow and stopped-flow experiments show that the first two processes, which occur in the 0.001-1.0-s time scale, can be correlated to the formation of two moles of tryptophanyl-adenylate per mole of dimeric enzyme. These two processes are reversible by adding PPi, as seen in the fluorimeter. The third process leads to a reaction product that can no longer reform ATP after addition of PPi and that represents tryptophanyl-ATP ester, as demonstrated by thin-layer chromatography. This compound has been previously shown to be formed by tryptophanyl-tRNA synthetase from E. coli [K. H. Muench (1969) Biochemistry 8, 4872-4879]. Its formation is accompanied by a fluorescence decrease which reaches a minimum in about 30 min. The nature of the fourth process depends on the reaction conditions employed. In sodium bicarbonate or potassium phosphate buffer, the fourth process corresponds to the non-enzymatic hydrolysis of tryptophanyl-ATP ester. This spontaneous hydrolysis competes with formation of the ester and limits its concentration. Eventually, the progressive exhaustion of ATP brings the fluorescence intensity of the reaction mixture back to its initial value. In contrast, in ammonium bicarbonate buffer the previous third process is no longer visible, as evidenced by the absence of a fluorescence decrease beyond the fast initial quenching linked to the formation of tryptophanyl-adenylate. Instead, a fluorescence increase is observed. However, unlike the fourth process seen in sodium bicarbonate buffer, the fluorescence increase in ammonium bicarbonate is much larger than the initial fluorescence decrease linked to adenylate formation, the final fluorescence greatly surpassing the starting fluorescence signal. The reaction product of this process is tryptophanamide, as evidenced by high-performance liquid chromatography. Tryptophanamide formation is faster than that of tryptophanyl-ATP ester and is enzyme-catalyzed with a Km of 1 mM for ammonia and a rate constant of 5.7 min-1 at pH 8.3, 25 degrees C. The affinity of tryptophanamide for the protein is too weak to allow the formation of a significant concentration of enzyme-product complex. Tryptophanamide is therefore released in the reaction medium and its concentration reaches that of the limiting substrate.
...
PMID:Tryptophanamide formation by Escherichia coli tryptophanyl-tRNA synthetase. 388 Dec 55

The interaction between mouse submaxillary gland renin and a statine-containing, iodinated substrate analog inhibitor was studied. The compound, 1 (Boc-His-Pro-Phe-(4-iodo)-Phe-Sta-Leu-Phe-NH2, Sta = (3S,4S)-4-amino-3-hydroxy-6-methyl-heptanoic acid), a statine-containing analog of the renin substrate octapeptide, was a competitive inhibitor of cleavage of synthetic tetradecapeptide renin substrate by mouse submaxillary gland renin, with a Ki of 6.2 x 10(-10) M (pH 7.2, 37 degrees C). Titration of the partial quenching of the tryptophan fluorescence of the enzyme by 1 revealed tight binding with a dissociation constant less than 3 nM and a binding stoichiometry of one mole 1 per mole enzyme. The time course of tight binding of 1 to mouse renin appeared to be fast, with kON greater than or equal to 1.3 x 10(6) s-1 M-1. The UV difference spectrum generated upon binding of 1 to mouse renin had two prominent features: a strong, broad band that had a minimum at 242 nm with delta epsilon (242) = -19,500 cm-1 M-1, and a triplet of enhanced bands centered at 286 nm with delta epsilon (286) about +1100 cm-1 M-1. The strong, broad, negative band was similar to the difference between the UV absorbance of 1 in methanol and in 0.1 M citrate phosphate pH 7.2. A structure-activity correlation for analogs of 1 showed some moieties of 1 that are important for potent inhibition of mouse renin. The inhibition data for these compounds versus human kidney renin suggested that the solution of the crystal structure of 1 bound to mouse renin will provide useful information for the design of inhibitors of human kidney renin.
...
PMID:Interaction of mouse submaxillary gland renin with a statine-containing, subnanomolar, competitive inhibitor. 391 10

<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>