| 10615944 |
Membrane glycoprotein PC-1 inhibition of insulin receptor function occurs via direct interaction with the receptor alpha-subunit |
10.2337/diabetes.49.1.13. |
Diabetes |
Membrane glycoprotein PC-1 inhibition of insulin receptor function occurs via direct interaction with the receptor alpha-subunit
Abstract
- Plasma cell membrane glycoprotein-1 (PC-1) inhibits insulin receptor (IR) tyrosine kinase activity and subsequent cellular signaling. PC-1 content is elevated in fibroblasts, muscle, and adipose tissue from insulin-resistant subjects, and its elevation correlates with in vivo insulin resistance. In vitro, when PC-1 is transfected and overexpressed in cultured cells, it inhibits IR tyrosine kinase activity. To determine the mechanism whereby PC-1 regulates the IR, we studied how PC-1 interacts with this protein. Overexpression of PC-1 in MCF-7 cells inhibited tyrosine kinase activity of the IR, but not of the IGF-I receptor. When the IR was immunocaptured by specific IR monoclonal antibodies, PC-1 was associated with this receptor. In contrast, after specific immunocapture, PC-1 was not associated with the IGF-I receptor. We next studied HTC cells that were overexpressing an IR alpha-subunit mutant. This IR mutant binds insulin but has a deletion in the tyrosine kinase regulatory domain located in amino acids 485-599. In contrast to normal IRs, PC-1 did not associate with this mutant and did not affect tyrosine kinase activity. To determine whether decreasing PC-1 expression would reverse the inhibition of tyrosine kinase activity, we treated MCF-7 cells overexpressing PC-1 with a monoclonal antibody to PC-1. This treatment decreased PC-1 levels; concomitantly, IR tyrosine kinase activity increased. In contrast, IGF-I receptor tyrosine kinase activity was not increased. These studies indicate, therefore, that PC-1 may inhibit the IR by interacting directly with a specific region in the IR alpha-subunit. These studies also raise the possibility that monoclonal antibodies to PC-1 could be a new treatment for insulin resistance.
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| 10619399 |
Genomic structure and transcriptional regulation of the human somatostatin receptor type 2. |
10.1016/s0303-7207(99)00161-6 |
Mol. Cell. Endocrinol. |
Genomic structure and transcriptional regulation of the human somatostatin receptor type 2.
Abstract
- Somatostatin exerts inhibitory effects on virtually all endocrine and exocrine secretions. The somatostatin receptor subtype 2 (sst2) acts as a critical molecule for growth hormone regulation and cell proliferation. We investigated the structure and regulation of the human sst2 gene. A genomic clone including the sst2 gene was isolated, 1.5 kb of the promoter was sequenced and putative transcription factor binding sites were identified. The transcription start site was located 93 nucleotides upstream of the translation start site. The nucleotide sequences of the complete gene and 0.5 kb of 3' region were determined. A possible polyadenylation signal was identified. Transcriptional regulation was investigated by transient transfections using various promoter fragments. A -1100 sst2 promoter directed significant levels of luciferase expression in GH4 rat pituitary cells and Skut1-B endometrium cells whereas only low activity was detected in JEG3 chorion carcinoma cells or COS-7 monkey kidney cells. A minimal -252 promoter allowed cell specific expression. We did not find any regulation of the sst2 promoter by somatostatin, forskolin, TRH, TPA, T3, and 17beta-estradiol. Glucocorticoids lead to a significant inhibition of sst2 promoter activity. Further mapping suggest a glucocorticoid-responsive element between -905 and -707 and between -252 and -163. These studies demonstrate the nature of the human sst2 gene and identify its 5' and 3' flanking regions. Furthermore, specific activity of the promoter and regulation by various hormones is demonstrated.
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| 10624986 |
The hemodynamic effects of endothelin receptor antagonism during a venous air infusion in dogs |
10.1097/00000539-200001000-00022. |
Anesth Analg |
The hemodynamic effects of endothelin receptor antagonism during a venous air infusion in dogs
Abstract
- Endothelin (ET) is involved in the humoral component of the vasoconstriction during pulmonary embolism. We examined the effects of selective ET receptor antagonists on the hemodynamic and respiratory changes and on serum thromboxane B2 (TXB2) levels, during a continuous venous air infusion (VAI) in anesthetized mongrel dogs. The VAI (0.2 mL x kg(-1) x min(-1)) was initiated 5 min after an injection of saline (controls, n = 7), 1 micromol of the selective ET(A) receptor antagonist JKC-301 (group A, n = 6), or 1 micromol of the selective ET(B) receptor antagonist BQ-788 (group B, n = 6). Hemodynamic evaluation was performed every 15 min of VAI, and blood samples were drawn for blood gas analysis and TXB2 determinations. The increase in pulmonary perfusion pressure after 30 min of VAI was attenuated in Group A compared with the controls and Group B (Group A = 7+/-1 mm Hg; Group B = 16+/-1 mm Hg; controls = 14+/-1 mm Hg; P < 0.05). Pulmonary vascular resistance showed a similar behavior. TXB2 concentrations increased after 60 min of VAI in the controls and in Group B, but not in Group A (controls = 48%; Group B = 104%; Group A = 18%; P < 0.05 for controls and Group B). Similar decreases in Pao2 and Sao2 were observed in the three groups. We conclude that antagonism of ET(A) receptors attenuates the hemodynamic changes and blunts the increase in thromboxane A2 production during a VAI in dogs.
We evaluated the effects of endothelin receptor antagonists during a venous air infusion in dogs. EndothelinA receptor antagonism attenuated the hemodynamic changes and blunted the increase in thromboxane A2 production in this setting.
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| 10644367 |
A role for SKIP in EBNA2 activation of CBF1-repressed promoters. |
10.1128/jvi.74.4.1939-1947.2000 |
J. Virol. |
A role for SKIP in EBNA2 activation of CBF1-repressed promoters.
Abstract
- EBNA2 is essential for Epstein-Barr virus (EBV) immortalization of B lymphocytes. EBNA2 functions as a transcriptional activator and targets responsive promoters through interaction with the cellular DNA binding protein CBF1. We have examined the mechanism whereby EBNA2 overcomes CBF1-mediated transcriptional repression. A yeast two-hybrid screen performed using CBF1 as the bait identified a protein, SKIP, which had not previously been recognized as a CBF1-associated protein. Protein-protein interaction assays demonstrated contacts between SKIP and the SMRT, CIR, Sin3A, and HDAC2 proteins of the CBF1 corepressor complex. Interestingly, EBNA2 also interacted with SKIP in glutathione S-transferase affinity and mammalian two-hybrid assays and colocalized with SKIP in immunofluorescence assays. Interaction with SKIP was not affected by mutation of EBNA2 conserved region 6, the CBF1 interaction region, but was abolished by mutation of conserved region 5. Mutation of conserved region 5 also severely impaired EBNA2 activation of a reporter containing CBF1 binding sites. Thus, interaction with both CBF1 and SKIP is necessary for efficient promoter activation by EBNA2. A model is presented in which EBNA2 competes with the SMRT-corepressor complex for contacts on SKIP and CBF1.
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| 10650072 |
Synthesis of the marine sponge cycloheptapeptide phakellistatin 5(1) |
10.1021/np990253+. |
J Nat Prod |
Synthesis of the marine sponge cycloheptapeptide phakellistatin 5(1)
Abstract
- Phakellistatin 5 (1), a constituent of The Federated States of Micronesia (Chuuk) marine sponge Phakellia costada, was synthesized by solution-phase and solid-phase techniques. Because the linear peptide bearing (R)-Asn resisted cyclization, the synthesis of this peptide was repeated using the PAL resin attachment proceeding from N-Fmoc-D-Asp-alpha-OCH(2)CH=CH(2). After addition of the final unit (Ala), the allyl ester was removed under neutral conditions with Pd(o) [P(C(6)H(5))(3)](4). Removal of the final Fmoc-protecting group and cyclization with PyAOP provided (R)-Asn-phakellistatin 5 (2) in 28% overall yield. The same synthetic route from (S)-Asp led to natural phakellistatin 5 (1) in 15% overall recovery. The solution-phase and solid-phase synthetic products derived from (S)-Asp were found to be chemically but not biologically identical with natural phakellistatin 5 (1). This important fact suggested that a trace, albeit highly cancer-cell growth inhibitory, constituent accompanied the natural product or that there is a subtle conformational difference between the synthetic and natural cyclic peptides.
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| 10650101 |
A new depsipeptide from the sacoglossan mollusk Elysia ornata and the green alga Bryopsis species |
10.1021/np990402o. |
J Nat Prod |
A new depsipeptide from the sacoglossan mollusk Elysia ornata and the green alga Bryopsis species
Abstract
- A new cyclic depsipeptide, kahalalide O (1), was isolated from the sacoglossan Elysia ornata and its algal diet Bryopsis sp. The structure was elucidated primarily by NMR and MS spectral methods, and the stereochemistry of the amino acid residues was determined by chiral HPLC and Marfey analyses. Unlike the related metabolite kahalalide F, which is in development as a potential anticancer agent, kahalalide O (1) was inactive in arresting the growth of P-388, A549, HT29, and MEL28 cancer cell lines in vitro.
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| 10651391 |
Quinupristin/dalfopristin: a review of its use in the management of serious gram-positive infections |
10.2165/00003495-199958060-00008. |
Drugs |
Quinupristin/dalfopristin: a review of its use in the management of serious gram-positive infections
Abstract
- Quinupristin/dalfopristin is the first parenteral streptogramin antibacterial agent, and is a 30:70 (w/w) ratio of 2 semisynthetic pristinamycin derivatives. The combination has inhibitory activity against a broad range of gram-positive bacteria including methicillin-resistant staphylococci, vancomycin-resistant Enterococcus faecium (VREF), drug-resistant Streptococcus pneumoniae, other streptococci, Clostridium perfringens and Peptostreptococcus spp. The combination also has good activity against selected gram-negative respiratory tract pathogens including Moraxella catarrhalis, Legioniella pneumophila and Mycoplasma pneumoniae. Quinupristin/dalfopristin has poor activity against E. faecalis. The combination is bactericidal against staphylococci and streptococci, although constitutive erythromycin resistance can affect its activity. As for many other agents, quinupristin/dalfopristin is generally bacteriostatic against E. faecium. In patients with methicillin-resistant S. aureus (MRSA) or VREF infections participating in prospective emergency-use trials, quinupristin/dalfopristin 7.5 mg/kg every 8 or 12 hours achieved clinical or bacteriological success in > or =64% of patients. Emergence of resistance to quinupristin/dalfopristin was uncommon (4% of patients) in those with VREF infections. Quinupristin/dalfopristin 7.5 mg/kg 8- or 12-hourly also achieved similar clinical success rates to comparator agents in patients with presumed gram-positive complicated skin and skin structure infections or nosocomial pneumonia (administered in combination with aztreoman) in 3 large multicentre randomised trials. Systemic adverse events associated with quinupristin/dalfopristin include gastrointestinal events (nausea, vomiting and diarrhoea), rash and pruritus. Myalgias and arthralgias also occur at an overall incidence of 1.3%, although higher rates (2.5 to 31%) have been reported in patients with multiple comorbidities. Venous events are common if the drug is administered via a peripheral line; however, several management options (e.g. use of central venous access, increased infusion volume) may help to minimise their occurrence. Hyperbilirubinaemia has been documented in 3.1% of quinupristin/dalfopristin recipients versus 1.3% of recipients of comparator agents. Quinupristin/dalfopristin inhibits cytochrome P450 3A4 and therefore has the potential to increase the plasma concentrations of substrates of this enzyme.
Quinupristin/dalfopristin, the first parenteral streptogramin, offers a unique spectrum of activity against multidrug-resistant gram-positive bacteria. In serious gram-positive infections for which there are other treatment options available, the spectrum of activity and efficacy of quinupristin/ dalfopristin should be weighed against its tolerability and drug interaction profile. However, in VREF or unresponsive MRSA infections, where few proven treatment options exist, quinupristin/dalfopristin should be considered as a treatment of choice for these seriously ill patients.
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| 10651828 |
Peptides with antimicrobial activity from four different families isolated from the skins of the North American frogs Rana luteiventris, Rana berlandieri and Rana pipiens |
10.1046/j.1432-1327.2000.01074.x. |
Eur J Biochem |
Peptides with antimicrobial activity from four different families isolated from the skins of the North American frogs Rana luteiventris, Rana berlandieri and Rana pipiens
Abstract
- The skins of frogs of the genus Rana synthesize a complex array of antimicrobial peptides that may be grouped into eight families on the basis of structural similarity. A total of 24 peptides with differential growth-inhibitory activity towards the Gram-positive bacterium Staphylococcus aureus, the Gram-negative bacterium Escherichia coli and the yeast Candida albicans were isolated from extracts of the skins of three closely related North American frogs, Rana luteiventris (spotted frog), Rana berlandieri (Rio Grande leopard frog) and Rana pipiens (Northern leopard frog). Structural characterization of the antimicrobial peptides demonstrated that they belonged to four of the known families: the brevinin-1 family, first identified in skin of the Asian frog Rana porosa brevipoda; the esculentin-2 family, first identified in the European frog Rana esculenta; the ranatuerin-2 family, first identified in the North American bullfrog Rana catesbeiana; and the temporin family, first identified in the European frog Rana temporaria. Peptides belonging to the brevinin-2, ranalexin, esculentin-1 and ranatuerin-1 families were not identified in the extracts. Despite the close phylogenetic relationship between the various species of Ranid frogs, the distribution and amino-acid sequences of the antimicrobial peptides produced by each species are highly variable and species-specific, suggesting that they may be valuable in taxonomic classification and molecular phylogenetic analysis.
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| 10655483 |
Identification of a transcriptional repressor related to the noncatalytic domain of histone deacetylases 4 and 5. |
10.1073/pnas.97.3.1056 |
Proc. Natl. Acad. Sci. U.S.A. |
Identification of a transcriptional repressor related to the noncatalytic domain of histone deacetylases 4 and 5.
Abstract
- Histone deacetylases (HDACs) are involved in regulating transcription by modifying the core histones of the nucleosome. To date, six HDACs have been identified in mammalian cells: the yeast RPD3 homologs HDAC1, 2, and 3 and the yeast HDA1 homologs HDAC4, 5, and 6. HDAC4 and HDAC5 contain a noncatalytic N-terminal domain. Herein, we report the identification of a protein HDRP (HDAC-related protein) that shares 50% identity in deduced amino acid sequence to the noncatalytic N-terminal domain of HDAC4 and 5. The steady-state levels of HDRP mRNA are high in human brain, heart, and skeletal muscle and low in the several other tissues. HDRP has an apparent molecular mass of approximately 75 kDa. HDRP does not possess intrinsic HDAC activity but forms complexes with both HDAC1 and HDAC3. HDRP represses both basal and activated transcription in transient transfection assays when tethered to DNA as a Gal4-fusion protein. HDAC inhibitors do not reverse transcriptional repression mediated by Gal4-HDRP. Thus, HDRP is a transcriptional repressor and can repress transcription in the presence of HDAC inhibitors.
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| 10669754 |
Sequestration and inhibition of Daxx-mediated transcriptional repression by PML. |
10.1128/mcb.20.5.1784-1796.2000 |
Mol. Cell. Biol. |
Sequestration and inhibition of Daxx-mediated transcriptional repression by PML.
Abstract
- PML fuses with retinoic acid receptor alpha (RARalpha) in the t(15;17) translocation that causes acute promyelocytic leukemia (APL). In addition to localizing diffusely throughout the nucleoplasm, PML mainly resides in discrete nuclear structures known as PML oncogenic domains (PODs), which are disrupted in APL and spinocellular ataxia cells. We isolated the Fas-binding protein Daxx as a PML-interacting protein in a yeast two-hybrid screen. Biochemical and immunofluorescence analyses reveal that Daxx is a nuclear protein that interacts and colocalizes with PML in the PODs. Reporter gene assay shows that Daxx drastically represses basal transcription, likely by recruiting histone deacetylases. PML, but not its oncogenic fusion PML-RARalpha, inhibits the repressor function of Daxx. In addition, SUMO-1 modification of PML is required for sequestration of Daxx to the PODs and for efficient inhibition of Daxx-mediated transcriptional repression. Consistently, Daxx is found at condensed chromatin in cells that lack PML. These data suggest that Daxx is a novel nuclear protein bearing transcriptional repressor activity that may be regulated by interaction with PML.
|
| 10673369 |
Marked increase in membranolytic selectivity of novel cyclic tachyplesins constrained with an antiparallel two-beta strand cystine knot framework |
10.1006/bbrc.1999.2035. |
Biochem Biophys Res Commun |
Marked increase in membranolytic selectivity of novel cyclic tachyplesins constrained with an antiparallel two-beta strand cystine knot framework
Abstract
- We have developed a highly constrained 18-residue cyclic peptide template based on the antimicrobial peptide tachyplesin-1 that features an end-to-end peptide backbone and a cystine knot-like motif with three evenly spaced disulfide bonds to cross-brace the antiparallel beta-strands and to approximate an amphiphatic "beta-tile"-like structure. Six beta-tile analogs were prepared to correlate different topological patterns with membranolytic specificity. Their conformations and antimicrobial and hemolytic activities were compared with tachyplesin-1 and the recently discovered Rhesus monkey theta defensin (RTD) which contains similar beta-tile structural elements. The beta-tile peptides and RTD retained broad spectrum antimicrobial activities. In general, they were less active than tachyplesin-1 in 10 tested organisms but their activity increased under high-salt (100 mM NaCl) rather than in low-salt conditions. The beta-tile peptides are highly nontoxic to human erythrocytes with EC(25) ranging from 600 to 4000 microM. Collectively, our results show that the design of a highly rigid peptide template is useful for further analog study to dissociate antimicrobial activity from cytotoxicity which would be helpful in discovering clinical applications for peptide antibiotics.
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| 10679990 |
Location of alkali metal binding sites in endothelin A selective receptor antagonists, cyclo(D-Trp-D-Asp-Pro-D-Val-Leu) and cyclo(D-Trp-D-Asp-Pro-D-Ile-Leu), from multistep collisionally activated decompositions |
10.1002/(SICI)1096-9888(200002)35:2<265::AID-JMS946>3.0.CO;2-#. |
J Mass Spectrom |
Location of alkali metal binding sites in endothelin A selective receptor antagonists, cyclo(D-Trp-D-Asp-Pro-D-Val-Leu) and cyclo(D-Trp-D-Asp-Pro-D-Ile-Leu), from multistep collisionally activated decompositions
Abstract
- We previously showed by using mass spectrometry that endothelin A selective receptor antagonists BQ123 and JKC301 form novel coordination compounds with sodium ions. This property may underlie the ability of an ET(A) antagonist to induce net tubular sodium reabsorption in the proximal tubule cells and reverse acute renal failure induced by severe ischemia. We have now defined the metal binding sites on BQ123 and JKC301 by subjecting the metal-containing peptides to multiple stages of collisionally activated decomposition (CAD) in an ion trap mass spectrometer. When submitted to low-energy CAD, the ring opens at the Asp-Pro amide bond. The metal ion, which bonds, inter alia, to the carbonyl oxygen of the proline residue, acts as a fixed charge site, and directs a charge-remote, sequence-specific fragmentation of the ring-opened peptide. Amino acid residues are sequentially cleaved from the C-terminal end, and the terminal aziridi structure moves one step toward the N-terminus with each C-terminal amino acid residue removed. These observations are the basis of a new method to sequence cyclic peptides. Amino acid residues are observed as sets of three ions, a*(n)PD, b*(n)PD and c*(n)PD where n is the number of amino acid residues in the peptide.
|
| 10684867 |
Neurons regulate extracellular levels of amyloid beta-protein via proteolysis by insulin-degrading enzyme. |
10.1523/jneurosci.20-05-01657.2000 |
J. Neurosci. |
Neurons regulate extracellular levels of amyloid beta-protein via proteolysis by insulin-degrading enzyme.
Abstract
- Progressive cerebral accumulation of amyloid beta-protein (Abeta) is an early and invariant feature of Alzheimer's disease. Little is known about how Abeta, after being secreted, is degraded and cleared from the extracellular space of the brain. Defective Abeta degradation could be a risk factor for the development of Alzheimer's disease in some subjects. We reported previously that microglial cells release substantial amounts of an Abeta-degrading protease that, after purification, is indistinguishable from insulin-degrading enzyme (IDE). Here we searched for and characterized a role for IDE in Abeta degradation by neurons, the principal cell type that produces Abeta. Whole cultures of differentiated pheochromocytoma (PC12) cells and primary rat cortical neurons actively degraded endogenously secreted Abeta via IDE. However, unlike that in microglia, IDE in differentiated neurons was not released but localized to the cell surface, as demonstrated by biotinylation. Undifferentiated PC12 cells released IDE into their medium, whereas after differentiation, IDE was cell associated but still degraded Abeta in the medium. Overexpression of IDE in mammalian cells markedly reduced the steady-state levels of extracellular Abeta(40) and Abeta(42), and the catalytic site mutation (E111Q) abolished this effect. We observed a novel membrane-associated form of IDE that is approximately 5 kDa larger than the known cytosolic form in a variety of cells, including differentiated PC12 cells. Our
Results support a principal role for membrane-associated and secreted IDE isoforms in the degradation and clearance of naturally secreted Abeta by neurons and microglia.
|
| 10690891 |
Human somatostatin receptor subtypes in acromegaly: distinct patterns of messenger ribonucleic acid expression and hormone suppression identify different tumoral phenotypes |
10.1210/jcem.85.2.6338. |
J Clin Endocrinol Metab |
Human somatostatin receptor subtypes in acromegaly: distinct patterns of messenger ribonucleic acid expression and hormone suppression identify different tumoral phenotypes
Abstract
- Recently, studies using somatostatin (SRIF) analogs preferential for either the SRIF receptor 2 (SSTR2) or the SSTR5 subtype demonstrated a variable suppression of GH and PRL release from GH-secreting human adenomas. These data suggested the concept of SSTR subtype specificity in such tumors. In the present study the quantitative expression of messenger ribonucleic acid (mRNA) for the 5 SSTR subtypes and the inhibitory effects of SRIF14; SRIF28; octreotide; the SSTR2-preferential analog, BIM-23197; and the SSTR5-preferential analog, BIM-23268, on GH and PRL secretion were analyzed in cells cultured from 15 acromegalic tumors. RT-PCR analysis revealed a consistent pattern of SSTR2 and SSTR5 mRNA expression. SSTR5 mRNA was expressed at a higher level (1052 +/- 405 pg/pg glyceraldehyde-3-phosphate dehydrogenase) than SSTR2 mRNA (100 +/- 30 pg/pg glyceraldehyde-3-phosphate dehydrogenase). However, only SSTR2 mRNA expression correlated with the degree of GH inhibition induced by SRIF14, SRIF28, and BIM-23197. The SSTR5-preferential compound inhibited GH release in only 7 of 15 cases. In cells cultured from the 10 mixed adenomas that secreted both GH and PRL, RT-PCR analysis revealed a consistent coexpression of SSTR5, SSTR2, and SSTR1 mRNA. In all cases SRIF14, SRIF28, and the SSTR5-preferential analog, BIM-23268, significantly suppressed PRL secretion, with a mean maximal inhibition of 48 +/- 4%. In contrast, the SSTR2-preferential analogs, BIM-23197 and octreotide, were effective in suppressing PRL in only 6 of 10 cases. In cells cultured from adenomas taken from patients partially responsive to the SRIF analog, octreotide, partial additivity in suppressing both GH and PRL secretion was observed when the SSTR2- and SSTR5-preferring analogs, BIM-23197 and BIM-23268, were tested in combination. Our data show a highly variable ratio of the SSTR2 and SSTR5 transcripts, according to tumors. The SSTR2-preferring compound consistently inhibits GH release, whereas the SSTR5-preferring compound is the main inhibitor of PRL secretion. When both drugs are combined, the partial additivity observed in mixed GH- plus PRL-secreting adenomas may be of interest in the therapeutic approach of such tumors.
|
| 10691702 |
New circulin macrocyclic polypeptides from Chassalia parvifolia |
10.1021/np990432r. |
J Nat Prod |
New circulin macrocyclic polypeptides from Chassalia parvifolia
Abstract
- Four new macrocyclic polypeptides were isolated and identified from an extract of the tropical tree Chassalia parvifolia. Circulins C-F are 29-30 amino acid cyclic peptides in which the entire primary amino acid chain is covalently cyclized via peptide bonds. Their structures were deduced from a combination of FABMS analyses, N-terminal Edman degradation, endoproteinase digestion, and amino acid analyses. All the peptides share a high degree of sequence homology and contain six cysteine residues forming three intramolecular disulfide bridges. Circulins C-F inhibited the cytopathic effects of in vitro HIV-1 infection with EC(50) values of 50-275 nM.
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