Home About us Contact
|
Home About us Contact | ||
|
|
||
Synthesized Peptides (synthesized + peptide)
Selected AbstractsIdentification of oligopeptides binding to peritoneal tumors of gastric cancerCANCER SCIENCE, Issue 10 2006Noriyuki Akita This is a report of in vivo intraperitoneal biopanning, and we successfully identified a novel peptide to target the multiple peritoneal tumors of gastric cancer. A phage display library was injected directly into the abdominal cavity of mice bearing peritoneal tumors of human gastric cancer, and phages associated with the tumors were subsequently reclaimed from isolated samples. The tumor-associated phages were amplified and the biopanning cycle was repeated five times to enrich for high affinity tumor-selective binding peptides. Finally, a tri-peptide motif, KLP, which showed homology with laminin 5 (a ligand for ,3,1 integrin), was identified as a binding peptide for peritoneal tumors of gastric cancer. Phage clones displaying the sequence KLP showed 64-fold higher binding to peritoneal tumors than control phage and were preferentially distributed in tumors rather than in normal organs after intraperitoneal injection into mice. In addition, the KLP phages were more likely to bind to cancer cells in malignant ascites derived from a patient with recurrent gastric cancer. Synthesized peptide containing the motif KLP (SWKLPPS) also showed a strong binding activity to peritoneal tumors without cancer growth effect. Liposomes conjugated with SWKLPPS peptide appeared significantly more often in tumors than control liposomes after intraperitoneal injection into mice. Furthermore, modification of liposomes with SWKLPPS peptide enhanced the antitumor activity of adriamycin on gastric cancer cells. The peptide motif KLP seems a potential targeting ligand for the treatment of peritoneal metastasis of gastric cancer. (Cancer Sci 2006; 97: 1075,1081) [source] Solid phase peptide synthesis on epoxy-bearing methacrylate monolithsJOURNAL OF PEPTIDE SCIENCE, Issue 12 2004E. Vlakh Abstract Monoliths based on a copolymer of glycidyl methacrylate (GMA) and ethylene dimethacrylate (EDMA) can be used directly as sorbents for affinity chromatography after solid phase peptide synthesis. The quality of the synthesized products, the amount of grown peptides on a support and the reproducibility of the process must be considered. A determination of the quantity of the introducing ,-Ala (and, consequently, the total amount of synthesized peptide) was carried out. Three peptides complementary to recombinant tissue plasminogen activator (t-PA) have been synthesized using Fmoc-chemistry on GMA-EDMA disks. The peptidyl ligands were analysed by amino acid analysis, ES-MS and HPLC methods. The affinity binding parameters were obtained from frontal elution data. The results were compared with those established for GMA-EDMA affinity sorbents formed by the immobilization of the same but separately synthesized and purified ligands. The immobilization on GMA-EDMA disks was realized using a one-step reaction between the amino groups of the synthetic ligand and the original epoxy groups of monolithic material. The affinity constants found for two kinds of sorbent did not vary significantly. Finally, the directly obtained affinity sorbents were tested for t-PA separation from a cellular supernatant. Copyright © 2004 European Peptide Society and John Wiley & Sons, Ltd. [source] Detection of tmRNA-mediated trans-translation products in Bacillus subtilisGENES TO CELLS, Issue 3 2002Ai Fujihara Background: Bacterial tmRNA (10Sa RNA) is involved in a trans -translation reaction, which contributes to the degradation of incompletely synthesized peptides and the recycling of stalled ribosomes. To investigate the physiological roles of this reaction in Bacillus subtilis, we devised a system for detecting the proteins that are subject to in vivo trans -translation. Results: The wild-type tmRNA gene (ssrA) in the genome was replaced by a variant ssrA encoding a tag-peptide sequence containing six histidine residues (His-tag) and two aspartic acids at the C-terminus. The His-tagged proteins that accumulated in the cells without degradation were fractionated by Ni2+ -NTA column and gel electrophoresis and were detected by Western blotting with an anti-His-tag antibody. The results showed that the trans -translation occurred more frequently at a high temperature (50 °C) than at a low temperature (37 °C). Two-dimensional (2D) gel electrophoresis of the products revealed many distinct spots, which represent specific target proteins for the trans -translation reaction. Furthermore, the 2D gel patterns of the products from cells cultured at high and low temperatures were apparently different. Several tagged proteins were identified by the N-terminal amino acid sequences of the products. Conclusion:Trans -translation occurs more frequently at high temperature than at low temperature, and different proteins are tagged at different temperatures. [source] Identification of a Peptide Sequence in Albumin that Potentiates Superoxide Production by MicrogliaJOURNAL OF NEUROCHEMISTRY, Issue 6 2000Yoichi Nakamura Abstract: Microglial activation has recently been recognized as acause of damage in various neurodegenerative diseases. A possible mechanismunderlying this damage is the activation of microglia by serum factors leakedthrough a disruption of the blood,brain barrier, which in turn triggermicroglial cell proliferation and the release of various substances toxic toneurons, such as superoxide (O2 - ). We recently reportedthat serum albumin enhanced O2 - producation in culturedrat microglia stimulated by phorbol ester. In the present report, we identifythe active site of this enhancement within the albumin molecule. We purifiedan active subfragment from trypsin-treated bovine serum albumin that wascomposed of 12-mer and 33-mer peptides connected by a disulfide bond. Thechemically synthesized 12-mer peptide showed activity within a concentrationrange (,10 -7M) equivalent to that of albumin. Theactivities of a series of synthesized peptides conclusively indicated that theminimum active sequence was Leu-His-Thr-Leu. The present study may shed lighton the mechanism of neuronal cell damage in various neurodegenerativediseases. [source] Elimination and exchange of trifluoroacetate counter-ion from cationic peptides: a critical evaluation of different approachesJOURNAL OF PEPTIDE SCIENCE, Issue 3 2008Stéphane Roux Abstract Most synthesized peptides are nowadays produced using solid-phase procedures. Due to cleavage and purification conditions, they are mainly obtained in the presence of trifluoroacetic acid (TFA) and, for cationic peptides, as trifluoroacetate (TF-acetate) salts. However, TF-acetate interferes with physicochemical characterizations using infrared spectroscopy and might significantly affect the in vivo studies. Thus, TF-acetate exchange by another counter-ion is often required. Up to now, the classical procedure has consisted of freeze-drying the peptide several times in the presence of an excess of a stronger acid than TFA (pKa ,0): generally HCl (pKa = , 7). This approach means that working at pH < 1 can induce peptide degradation. We therefore tested three different approaches to exchange the tightly bound TF-acetate counter-ion from the dicationic octapeptide lanreotide: (i) reverse-phase HPLC, (ii) ion-exchange resin, and (iii) deprotonation/reprotonation cycle of the amino groups. The first two approaches allow the partial to almost complete exchange of the TF-acetate counter-ion by another ion from an acid weaker than TFA, such as acetic acid (pKa = 4.5), and the third requires a basic solution that permits the complete removal of TF-acetate counter-ion. The efficiency of these three procedures was tested and compared by using different analytical techniques such as 19F-NMR, 1H-NMR and attenuated total reflectance Fourier transformed infrared spectroscopy (ATR FT-IR). We also show that ATR-IR can be used to monitor the TFA removal. The counter-ion exchange procedures described in this study are easy to carry out, fast, harmless and reproducible. Moreover, two of them offer the very interesting possibility of exchanging the initial TF-acetate by any other counter-ion. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd. [source] Emerging views on tmRNA-mediated protein tagging and ribosome rescueMOLECULAR MICROBIOLOGY, Issue 4 2001Reynald Gillet Transfer- messenger RNA (tmRNA), also known as SsrA or 10Sa RNA, is a bacterial ribonucleic acid that recycles 70S ribosomes stalled on problematic messenger RNAs (mRNAs) and also contributes to the degradation of incompletely synthesized peptides. tmRNA acts initially as transfer RNA (tRNA), being aminoacylated at its 3,-end by alanyl-tRNA synthetase, to add alanine to the stalled polypeptide chain. Resumption of translation ensues not on the mRNA on which the ribosomes were stalled but at an internal position in tmRNA. Termination soon occurs, tmRNA recruiting the appropriate termination factors allowing the release of the tagged protein that is subsequently recognized and degraded by specific cytoplasmic and periplasmic proteases, and permits ribosome recycling. Recent data suggest that tmRNA tags bacterial proteins in three other instances; when ribosomes stall at internal sites; during ,readthrough' of canonical termination codons; and when ribosomes are at the termination codon of intact messages. The importance of bacterial tmRNAs for survival, growth under stress, and pathogenesis is also discussed. Recent in vivo and in vitro studies have identified novel ligands of tmRNA. Based on the available experimental evidences, an updated model of tmRNA mediated protein tagging and ribosome rescue in bacteria is presented. [source] Isolation and characterization of cytotoxic small peptides, ,-casecidins, from bovine ,s1-casein digested with bovine trypsinANIMAL SCIENCE JOURNAL, Issue 5 2003Hajime OTANI ABSTRACT Cytotoxic peptides were isolated from a trypsin digest of bovine ,s1-casein by a combination of ion-exchange column chromatography and reversed-phase high-performance liquid chromatography as an indicator of cytotoxicity toward mouse spleen cells. Amino acids of the isolated peptides were sequenced as arginyl-proly-lysine, leucyl-lysyl-lysine and tyrosyl-lysine, being compatible with sequences 1,3, 101,103 and 104,105 of bovine ,s1-casein, respectively. The isolated peptides displayed cytotoxicity toward healthy mouse T and B cells and human leukemic T and B cell lines in a commercially available serum-free medium for lymphocytes, Celgrosser-P, and were named ,-casecidins. Similar cytotoxicity was confirmed in chemically synthesized peptides corresponding to sequences 1,3, 101,103 and 100,105 of bovine ,s1-casein. The cytotoxicity induced by ,-casecidins was concluded to be because of necrosis, and was diminished in the presence of bovine serum albumin. [source] | ||||||||||