| 1849131 |
Anantin--a peptide antagonist of the atrial natriuretic factor (ANF). I. Producing organism, fermentation, isolation and biological activity |
10.7164/antibiotics.44.164. |
J Antibiot (Tokyo) |
Anantin--a peptide antagonist of the atrial natriuretic factor (ANF). I. Producing organism, fermentation, isolation and biological activity
Abstract
- Anantin, a peptide binding to the receptor of the atrial natriuretic factor (ANF) was isolated from a strain of Streptomyces coerulescens. The molecule consists of 17 natural L-amino acids which form a peptidic ring system. It has a MW of 1,871.0. The chemical composition is C90H111N21O24. The compound was found to bind competitively to ANF-receptors from bovine adrenal cortex (Kd = 0.61 microM). Furthermore, it dose-dependently inhibited the ANF-induced intracellular cyclic guanosine monophosphate accumulation in bovine aorta smooth muscle cells. At the same concentration no agonistic effects were detectable in these cells. Thus, anantin is considered to be the first microbially produced antagonist of the cardiac hormone, ANF.
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| 1849260 |
Molecular cloning of a cDNA of a camptothecin-resistant human DNA topoisomerase I and identification of mutation sites |
10.1093/nar/19.1.69. |
Nucleic Acids Res |
Molecular cloning of a cDNA of a camptothecin-resistant human DNA topoisomerase I and identification of mutation sites
Abstract
- Camptothecin (CPT), a plant alkaloid with antitumor activity, is a specific inhibitor of eukaryotic DNA topoisomerase I. We have previously isolated and characterized a CPT-resistant topoisomerase I isolated from a CPT-resistant human leukemia cell line, CPT-K5. cDNA clones of topoisomerase I were isolated from the CPT-resistant and the parental CPT-sensitive cell lines, respectively. Sequencing of the clones identified two mutations in the cDNA isolated from the resistant cells, which cause amino acid changes from aspartic acid to glycine at residues 533 and 583 of the parental topoisomerase I. When the CPT-K5 topoisomerase I was expressed in E. coli as a fusion protein with Staphylococcal Protein A fragment, the activity was resistant to CPT at a dose level up to 125 microM, whereas the parental fusion protein was sensitive to CPT as low as 1 microM. The resistance index (greater than 125) of the CPT-K5 fusion topoisomerase I is similar to that of the native CPT-K5 topoisomerase I. These results indicate that either or both of the two amino acid changes identified in the mutant enzyme is responsible for the resistance to CPT.
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| 1851751 |
Structure of the human type I DNA topoisomerase gene. |
10.1016/s0021-9258(18)92864-4 |
J. Biol. Chem. |
Structure of the human type I DNA topoisomerase gene.
Abstract
- We describe the molecular organization of the human gene coding for type I DNA topoisomerase. The coding sequence is split into 21 exons distributed over at least 85 kilobase pairs (kb) of human genomic DNA. The sizes of the 20 introns vary widely between 0.2 and at least 30 kb and all contain the sequence elements known to be required for pre-mRNA splicing. Several of the intron sequences separate exons encoding parts of the enzyme that are highly conserved between human and yeast suggesting that at least some of the exons may code for individual, structurally, or functionally important domains of the enzyme. We also describe the promoter sequence of the human topoisomerase I gene and show that it is composed of distinct functional elements.
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| 1862749 |
Different inhibitory actions of cyclosporine A and cyclosporine A-acetate on lipopolysaccharide-, interleukin 1-, 1,25 dihydroxyvitamin D3- and parathyroid hormone-stimulated calcium and lysosomal enzyme release from mouse calvaria in vitro |
10.1007/BF01980893. |
Agents Actions |
Different inhibitory actions of cyclosporine A and cyclosporine A-acetate on lipopolysaccharide-, interleukin 1-, 1,25 dihydroxyvitamin D3- and parathyroid hormone-stimulated calcium and lysosomal enzyme release from mouse calvaria in vitro
Abstract
- The effects of cyclosporine A (CsA) and one of its immunologically inactive structural analogues, cyclosporine A acetate (CsA-A) on lipopolysaccharide (LPS)-, interleukin-1 (IL-1)-, 1,25 dihydroxyvitamin D3 (1,25 D3)- and parathyroid hormone (PTH)-stimulated bone resorption were tested in mouse calvaria cultures. The release of calcium and a lysosomal enzyme, N-acetylglycosaminidase (NAG) was determined after 3 days of culture. All bone resorbing agents potently stimulated calcium and NAG release. At therapeutic concentration levels of 0.1 and 1.0 micrograms/ml, the immunologically active CsA was significantly more potent than the inactive CsA-A against LPS- and 1,25 D3-induced calcium and NAG release. The inhibition by both cyclosporines of IL-1 and PTH stimulated calcium release was not significantly different. CsA was however more potent than CsA-A against IL-1 stimulated NAG release. PTH-stimulated NAG release was not inhibited by CsA or CsA-A. These findings suggest that both cyclosporines interfere with more than one mechanism of activation of bone resorption. The specific effect of CsA against LPS and 1,25 D3 may be related to its known inhibition of immune cell derived cytokine expression.
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| 1865337 |
Synthesis of a fluorescent derivative of cyclosporin A for high-performance liquid chromatography analysis |
10.1002/jps.2600800416. |
J Pharm Sci |
Synthesis of a fluorescent derivative of cyclosporin A for high-performance liquid chromatography analysis
Abstract
- A direct assay method for use in studies of cyclosporin binding must be highly sensitive and selective since it must be capable of measuring the concentrations encountered in the protein-free matrix. The failure of current HPLC methods to achieve the sensitivity required for binding studies may be attributed to the use of UV detection, which relies on the relatively weak end-absorption of cyclosporin A. A method involving fluorescence derivatization was sought with the aim of increasing HPLC assay sensitivity. A method is described for producing a fluorescent derivative of cyclosporin A, a compound which has no functional groups which are easily derivatized. However, intramolecular rearrangement of cyclosporin A to form its structural isomer, isocyclosporin A, exposes a secondary amine which can be reacted with dansyl chloride to produce a fluorescent derivative. This two-step derivatization procedure was used as the basis of an HPLC fluorescence assay. Although this assay is not sufficiently sensitive to measure concentrations encountered in the protein-free matrix during plasma binding studies, the method does point to the possible development of a more sensitive assay using a derivatizing reagent other than dansyl chloride.
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| 1872611 |
Purification and amino acid sequence of lactocin S, a bacteriocin produced by Lactobacillus sake L45 |
10.1128/aem.57.6.1829-1834.1991. |
Appl Environ Microbiol |
Purification and amino acid sequence of lactocin S, a bacteriocin produced by Lactobacillus sake L45
Abstract
- Lactocin S, a bacteriocin produced by Lactobacillus sake L45, has been purified to homogeneity by ion exchange, hydrophobic interaction and reverse-phase chromatography, and gel filtration. The purification resulted in approximately a 40,000-fold increase in the specific activity of lactocin S and enabled the determination of a major part of the amino acid sequence. Judging from the amino acid composition, lactocin S contained approximately 33 amino acid residues, of which about 50% were the nonpolar amino acids alanine, valine, and leucine. Amino acids were not detected upon direct N-terminal sequencing, indicating that the N-terminal amino acid was blocked. By cyanogen bromide cleavage at an internal methionine, the sequence of the 25 amino acids (including the methionine at the cleavage site) in the C-terminal part of the molecule was determined. The sequence was Met-Glu-Leu-Leu-Pro-Thr-Ala-Ala-Val-Leu-Tyr-Xaa-Asp-Val-Ala-Gly-Xaa-Phe- Lys-Tyr-Xaa-Ala-Lys-His-His, where Xaa represents unidentified residues. It is likely that the unidentified residues are modified forms of cysteine or amino acids associated with cysteine, since two cysteic acids per lactocin S molecule were found upon performic acid oxidation of lactocin S. The sequence was unique when compared to the SWISS-PROT data bank.
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| 1874714 |
In vitro biosynthesis of Thr2,Leu5,D-Hiv8,Leu10-cyclosporin, a cyclosporin-related peptolide, with immunosuppressive activity by a multienzyme polypeptide |
None |
J Biol Chem |
In vitro biosynthesis of Thr2,Leu5,D-Hiv8,Leu10-cyclosporin, a cyclosporin-related peptolide, with immunosuppressive activity by a multienzyme polypeptide
Abstract
- A new cyclic peptolide (SDZ 214-103), which is produced by the fungus Cylindrotrichumoligospermum (Corda) BONORDEN (Dreyfuss, M. M., Schreier, M. H., Tscherter, H., and Wenger, R. (June 15, 1988) European Patent Application 0 296 123 A2) and is closely related to cyclosporin A (CyA), has as the main structural difference D-2-hydroxyisovaleric acid in ester linkage at position 8 instead of D-alanine in the cyclosporins. This peptolide exerts similar biological activities to CyA. We were able to prepare an enzyme fraction of crude extracts of the mycelium, which is capable of synthesizing the peptolide with consumption of the constitutive amino acids, D-2-hydroxyisovaleric acid, ATP, and S-adenosyl-L-methionine. The in vitro product co-chromatographs with authentic peptolide on thin layer chromatography and high performance liquid chromatography and shows similar immunosuppressive activity in vitro. The enzyme does not synthesize CyA, whereas cyclosporin synthetase does not synthesize the peptolide. Peptolide synthetase has a high molecular weight (in the same range as cyclosporin synthetase) and also does not appear to be glycosylated. The enzyme cross-reacts with antibodies directed specifically against cyclosporin synthetase.
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| 1876038 |
Chronic cocaine-induced supersensitivity id blocked by co-treatment with the peptide cyclo-(Leu-Gly) |
None |
NIDA Res Monogr |
Chronic cocaine-induced supersensitivity id blocked by co-treatment with the peptide cyclo-(Leu-Gly)
Abstract
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| 1890161 |
A mutation in the tyrosine kinase domain of the insulin receptor associated with insulin resistance in an obese woman |
10.1210/jcem-73-4-894. |
J Clin Endocrinol Metab |
A mutation in the tyrosine kinase domain of the insulin receptor associated with insulin resistance in an obese woman
Abstract
- Insulin resistance is frequently associated with acanthosis nigricans and hyperandrogenism. In patients with type A insulin resistance, this has been shown to be due to genetic defects in insulin receptor function. However, other patients with a similar clinical syndrome have been reported to have a variant of this syndrome, in which assays of insulin receptor function were normal. We have sequenced a portion of the insulin receptor gene in one such patient, a 29-yr-old woman with obesity and insulin resistance. The patient is heterozygous for a mutation substituting isoleucine for methionine at position 1153. Met1153 is located in the intracellular domain of the receptor near the cluster of tyrosine phosphorylation sites at positions 1158, 1162, and 1163. Studies of the mutant receptor expressed in NIH-3T3 cells demonstrated that the Ile1153-mutation impairs the ability of insulin to stimulate autophosphorylation of solubilized insulin receptors. In addition, the mutation impairs the ability of insulin to stimulate receptor tyrosine kinase activity to phosphorylate an artificial substrate [poly(Glu-Tyr)]. It seems likely that this defect in receptor tyrosine kinase activity explains the defect in the ability of the patient's insulin receptors to mediate insulin action in vivo. Furthermore, this patient provides a paradigm in which genetic factors act in concert with other risk factors, such as obesity, to cause clinically important insulin resistance.
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| 1893911 |
Bradykinin agonist activity of a novel, potent oxytocin antagonist |
10.1016/0014-2999(91)90435-s. |
Eur J Pharmacol |
Bradykinin agonist activity of a novel, potent oxytocin antagonist
Abstract
- From a series of potent cyclic hexapeptide oxytocin (OT) antagonists, a compound that exhibited significant bradykinin (BK) agonist activity was identified. L-366,811 (cycloL-proline-D-tryptophan-L-isoleucine-D-pipecolic acid-L-piperazine-2-carboxylic acid-N-Me-D-phenylalanine) stimulated phosphatidylinositol (PI) turnover in rat uterine slices in vitro (approximately EC50, 2 microM) with a maximal effect (15-fold increase over basal) greater than that obtained for either BK or OT. L-366,811 also elicited dose-related contractions of the isolated rat uterus, producing measurable effects at 100 nM. Several other equally potent OT antagonists from the cyclic hexapeptide structural class were either less potent or inactive as activators of uterine PI turnover or contractility. The stimulatory effects of L-366,811 on uterine PI turnover and contractions were blocked by BK antagonists but not by an arginine vasopressin (AVP)/OT antagonist. In radioligand binding studies, L-366,811 exhibited moderate affinity (IC50, 360 nM) for the 3HBK binding site in rat uterus, consistent with its potency in the functional models. These results indicate that L-366,811 exhibits BK agonist activity in rat uterus in vitro.
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| 1903001 |
The uptake of the cyanobacterial hepatotoxin microcystin by isolated rat hepatocytes |
10.1016/0041-0101(91)90038-s. |
Toxicon |
The uptake of the cyanobacterial hepatotoxin microcystin by isolated rat hepatocytes
Abstract
- Microcystin-YM a cyclic heptapeptide hepatotoxin isolated from the cyanobacterium Microcystis aeruginosa was radiolabeled with 125I, and used to investigate the uptake of the toxin by freshly isolated rat hepatocytes. The uptake was temperature dependent with apparent activation energy of 18 kcal/mole (77 kJ/mole) for the initial rate of uptake. Uptake of non-toxic (10-20 nM) doses of microcystin by hepatocytes continued with time, the intracellular to extracellular distribution ratio for the toxin was 70 at 60 min for 10(6) cells/ml. Uptake of higher doses of microcystin (100 nM and more) stopped when the cells blebbed: a toxic response of hepatocytes to microcystin. Uptake of microcystin by hepatocytes was inhibited 70-80% by the addition of 10 microM sodium deoxycholate or bromsulphthlein, compounds that protect hepatocytes from the toxic effects of microcystin.
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| 1905540 |
Identification of amino acid substitutions in the lipopeptide surfactin using 2D NMR spectroscopy |
10.1016/0006-291x(91)90637-m. |
Biochem Biophys Res Commun |
Identification of amino acid substitutions in the lipopeptide surfactin using 2D NMR spectroscopy
Abstract
- It is generally accepted that surfactin, being produced by various Bacillus subtilis strains, is a cyclic lipopeptide built from the heptapeptide L-Glu-L-Leu-D-Leu-L-Val-L-Asp-D-Leu-L-Leu and a beta-hydroxy fatty acid with variable chain length of 13 - 15 carbon atoms. We investigated surfactin from Bacillus subtilis ATCC 21332 and OKB 105, dissolved in pyridine and methanol, with two-dimensional H NMR spectroscopy. In the NH-fingerprint region, 21 well resolved cross peaks are observed instead of the expected seven cross peaks for the given heptapeptide. We were able to assign all proton signals to 21 amino acids, to identify three heptapeptides, and thus to prove the existence of structural analogues of surfactin. In the major fraction A, the peptide sequence is as given above. In fractions B and C, the C-terminal leucine is replaced by valine and isoleucine, respectively.
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| 1907412 |
Isolation of two toxic heptapeptide microcystins from an axenic strain of Microcystis aeruginosa, K-139 |
10.1016/0041-0101(91)90022-j. |
Toxicon |
Isolation of two toxic heptapeptide microcystins from an axenic strain of Microcystis aeruginosa, K-139
Abstract
- Two toxic heptapeptides were isolated from an axenic Microcystis aeruginosa strain, K-139. Using mainly a non-destructive NMR method, we determined the structure of the major toxin to be 7-desmethylmicrocystin LR which lacks an N-methyl group of the dehydroalanine moiety of microcystin LR. The minor toxin was deduced to be 3,7-didesmethylmicrocystin LR. The chromatographic and NMR analyses of 7-desmethylmicrocystin LR were compared with those of 3-desmethylmicrocystin LR.
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| 1917291 |
Conformation of MeAla6-cyclosporin A by NMR. Relationship of sidechain orientation of the MeBmt-1, MeLeu-9, and MeLeu-10 residues to immunosuppressive activity |
None |
Int J Pept Protein Res |
Conformation of MeAla6-cyclosporin A by NMR. Relationship of sidechain orientation of the MeBmt-1, MeLeu-9, and MeLeu-10 residues to immunosuppressive activity
Abstract
- MeAla6-cyclosporin A (MeAla6-CsA) is a unique CsA analog that shows weak immunosuppressive activity and yet binds strongly to the proposed cytosolic protein receptor, cyclophilin (CyP). Preliminary 1H NMR data showed significant chemical shift differences between spectra of MeAla6-CsA and CsA, suggesting different preferred conformations. A more detailed study, however, revealed that the backbone conformations of the two molecules are essentially identical, and that the differences can be accounted for, principally, by the sidechain motions of the MeBmt-1, MeLeu-9, and -10 residues. ROE and coupling constant data show that in MeAla6-CsA, the preferred chi 1 rotamers for MeLeu-9 and -10 are + 180 degrees (T), whereas in CsA there is a more even distribution of rotamer populations for MeLeu-10, and a preferred -60 degrees (G-) chi 1 rotamer for MeLeu-9. Similar data argue that the sidechain of MeBmt-1 is more restricted in its motion in MeAla-CsA than in CsA. Temperature studies suggest that these preferred rotamers for MeAla6-CsA may increase the stability of the hydrogen bond between NH(7) and CO(11), but prevent particular residues, especially the essential MeBmt-1 sidechain, from adopting orientations required to elicit immunosuppressive activity. The significant changes observed in the preferred orientations for the sidechains of the MeBmt-1, MeLeu-9, and MeLeu-10 residues in MeAla6-CsA argue that the particular orientations which they assume in CsA are not essential for cyclophilin binding.
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| 1920361 |
Development of a small RGD peptide fibrinogen receptor antagonist with potent antiaggregatory activity in vitro |
10.1021/jm00114a022. |
J Med Chem |
Development of a small RGD peptide fibrinogen receptor antagonist with potent antiaggregatory activity in vitro
Abstract
- The development of potent antithrombotic agents from the fibrinogen platelet receptor binding sequences Fg-alpha 572-575 -Arg-Gly-Asp-Ser- and Fg-gamma 400-411 -HHLGGAKQAGDV, believed to be a cryptic RGD-type sequence, is described. The tetrapeptide Ac-RGDS-NH2 itself is capable of inhibiting platelet aggregation in vitro at high concentrations, IC50 91.3 +/- 0.1 microM in vitro antiaggregatory activity employing dog platelet rich plasma (PRP)/ADP, due to low platelet fibrinogen receptor affinity, Ki 2.9 +/- 1.9 microM (purified, reconstituted human platelet GPIIb/IIIa), relative to fibrinogen, Ki 38.0 +/- 6.0 nM. The peptide is also unstable to plasma, suffering total loss of in vitro activity upon incubation in PRP for 3 h (T1/2 90 min). Only modest improvements in potency were achieved with linear analogues of Ac-RGDS-NH2, while dramatic results were achieved with cyclic analogues, culminating in the cyclic disulfide Ac-cyclo-S,S-Cys-(N alpha-Me)Arg-Gly-Asp-Pen-NH2 (SK&F 106760) with improved plasma stability (100% activity after 3 h), affinity (Ki 58 +/- 20 nM purified human receptor), and potency (IC50 0.36 +/- 0.4 microM dog PRP/ADP). The affinity of this peptide is 2 orders of magnitude greater than that of Ac-RGDS-NH2. The affinity of the analogue is also comparable to fibrinogen. This peptide constitutes a first potent small peptide entry into the class of novel antithrombotic agents called fibrinogen receptor antagonists.
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