Embedding the Amyloid β-Peptide Sequence in Green Fluorescent Protein Inhibits Aβ Oligomerization

ChemBioChem ◽  
2007 ◽  
Vol 8 (9) ◽  
pp. 985-988 ◽  
Author(s):  
Tsuyoshi Takahashi ◽  
Kenichi Ohta ◽  
Hisakazu Mihara
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Amyloid Β ◽  
Peptide Sequence ◽  
Amyloid Β Peptide ◽  
Green Fluorescent ◽  
Aβ Oligomerization

Disulphide-less crotamine is effective for formation of DNA–peptide complex but is unable to improve bovine embryo transfection

Zygote ◽  
2019 ◽  
Vol 28 (1) ◽  
pp. 72-79
Author(s):  
Vicente J.F. Freitas ◽  
Iana S. Campelo ◽  
Mirelly M.A.S. Silva ◽  
Camila M. Cavalcanti ◽  
Dárcio I.A. Teixeira ◽  
...  
Keyword(s):  
Gene Expression ◽  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Peptide Sequence ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Gfp Expression ◽  
Green Fluorescent ◽  
Exogenous Gene

SummaryThis study aimed to investigate the ability of disulphide-less crotamine (dLCr) to complex DNA and to evaluate whether the DNA–dLCr complex is capable of improving transfection in bovine embryos. Three experiments were performed to: (i) evaluate the formation and stability of the DNA–dLCr complex; (ii) assess the dLCr embryotoxicity by exposure of bovine embryos to dLCr; and (iii) assess the efficiency of bovine embryo transfection after microinjection of the DNA–dLCr complex or green fluorescent protein (GFP) plasmid alone (control). DNA complexation by dLCr after 30 min of incubation at 1:100 and 1:50 proportions presented higher efficiency (P < 0.05) than the two controls: native crotamine (NCr) 1:10 and lipofectamine. There was no difference between DNA–dLCr 1:25 and the controls. The DNA–dLCr complexation was evaluated at different proportions and times. In all, at least half of maximum complexation was achieved within the initial 30 min. No embryotoxicity of dLCr was verified after exposure of in vitro fertilized embryos to different concentrations of the peptide. The effectiveness of dLCr to improve exogenous gene expression was evaluated by microinjection of the DNA–dLCr complex into in vitro fertilized zygotes, followed by verification of both embryo development and GFP expression. From embryos microinjected with DNA only, 4.6% and 2.8% expressed the GFP transgene at day 5 and day 7, respectively. The DNA–dLCr complex did not increase the number of GFP-positive embryos. In conclusion, dLCr forms a complex with DNA and its application in in vitro culture is possible. However, the dLCr peptide sequence should be redesigned to improve GFP expression.

scholarly journals Quick Delivery of Aldehyde Dehydrogenase into Yeast Vacuoles by QRPL Peptide Sequence on Carboxypeptidase Y

2020 ◽  
Author(s):  
Dong Jun Park ◽  
Ngoc-Tu Nguyen ◽  
Ji Hun Kim ◽  
Ngoc-Han Nguyen ◽  
Sunchang Kim ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Aldehyde Dehydrogenase ◽  
Signal Peptide ◽  
Fluorescent Protein ◽  
Peptide Sequence ◽  
Carboxypeptidase Y ◽  
Signal Peptide Sequence ◽  
Green Fluorescent ◽  
Vacuolar Protein ◽  
Vacuolar Targeting

Abstract Background The signal peptide sequence is known to increase transport efficiency to organelles in eukaryotic cells. In this study, we focus on the signal peptide of the vacuolar protein for vacuolar targeting. Results The signal peptide sequence QRPL of carboxypeptidase Y (CPY), a vacuolar protein, was inserted inside the green fluorescent protein (GFP) that does not locate in vacuole for vacuolar targeting. The protein location was then confirmed by confocal microscopy. Fascinatingly, the green fluorescent protein that contains QRPL inside the sequence could be expressed faster than its natural form (within 1 hour after induction). In addition, aldehyde dehydrogenase 6 (ALD6), a cytosolic protein has engineered the sequence with QRPL to be transported to the vacuole. The aldehyde removal activity of ALD6 protein in the recombinant yeast was then analyzed by measuring the luminescent intensity in Vibrio fischeri . Conclusions In summary, the signal peptide QRPL could be used not only to transport target proteins accurately to vacuole but also to improve the protein activity, as well as to shorten the induction time.

scholarly journals Identification of the amino acid sequence that targets peroxiredoxin 6 to lysosome-like structures of lung epithelial cells

2009 ◽  
Vol 297 (5) ◽  
pp. L871-L880 ◽  
Author(s):  
Elena M. Sorokina ◽  
Sheldon I. Feinstein ◽  
Tatyana N. Milovanova ◽  
Aron B. Fisher
Keyword(s):  
Amino Acids ◽  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Terminal Region ◽  
Peptide Sequence ◽  
Lamellar Bodies ◽  
Peroxiredoxin 6 ◽  
Lung Epithelial ◽  
Green Fluorescent ◽  
Acidic Organelles

Peroxiredoxin 6 (Prdx6), an enzyme with glutathione peroxidase and PLA2 (aiPLA2) activities, is highly expressed in respiratory epithelium, where it participates in phospholipid turnover and antioxidant defense. Prdx6 has been localized by immunocytochemistry and subcellular fractionation to acidic organelles (lung lamellar bodies and lysosomes) and cytosol. On the basis of their pH optima, we have postulated that protein subcellular localization determines the balance between the two activities of Prdx6. Using green fluorescent protein-labeled protein expression in alveolar epithelial cell lines, we showed Prdx6 localization to organellar structures resembling lamellar bodies in mouse lung epithelial (MLE-12) cells and lysosomes in A549 cells. Localization within lamellar bodies/lysosomes was in the luminal compartment. Targeting to lysosome-like organelles was abolished by the deletion of amino acids 31–40 from the Prdx6 NH2-terminal region; deletion of the COOH-terminal region had no effect. A green fluorescent protein-labeled peptide containing only amino acids 31–40 showed lysosomal targeting that was abolished by mutation of S32 or G34 within the peptide. Studies with mutated protein indicated that lipid binding was not necessary for Prdx6 targeting. This peptide sequence has no homology to known organellar targeting motifs. These studies indicate that the localization of Prdx6 in acidic organelles and consequent PLA2 activity depend on a novel 10-aa peptide located at positions 31–40 of the protein.

Sign in / Sign up

Export Citation Format

Share Document