ABSTRACT
Ricin is a highly toxic type 2 ribosome-inactivating protein (RIP) which is extracted from the seeds of castor beans. Ricin is considered a potential bioterror agent and no effective antidote for ricin exists so far. In this study, by structural modification of a retrograde transport blocker Retro-2, a series of novel compounds were obtained. The primary screen revealed that compound has an improved anti-ricin activity compare to positive control. pre-exposure evaluation in Madin-Darby Canine Kidney (MDCK) cells demonstrated that is a powerful anti-ricin compound with an EC of 41.05 nmol/L against one LC (lethal concentration, 5.56 ng/mL) of ricin. Further studies surprisingly indicated that confers post-exposure activity against ricin intoxication. An study showed that 1 h post-exposure administration of can improve the survival rate as well as delay the death of ricin-intoxicated mice. A drug combination of with monoclonal antibody mAb4C13 rescued mice from one LD (lethal dose) ricin challenge and the survival rate of tested animals is 100%. These results represent, for the first time, indication that small molecule retrograde transport blocker confers both and post-exposure protection against ricin and therefore provides a promising candidate for the development of anti-ricin medicines.
ABSTRACT
Objective To study the effect and mechanism of Momordica anti-HIV protein of 30 ku (MAP30) on the migration of bladder cancer.Methods The IC50 of human bladder cancer 5637 and T24 cells was calculated by methyl thiazolyl tetrazolium (MTT) method.The migration ability of these two cells was evaluated by scratch migration test and Transwell cell migration test.The expression of migrating proteins such as matrix metalloproteinases (MMPs) and adhesion molecule N-cadherin were compared by Western blot.Results Scratch migration test:there were significant differences in migration rates of 5637 cells at 8 h and 22 h (P ＜ 0.05).There were significant differences in migration rates of T24 cells at 22 h (P ＜ 0.05),but no significant differences in migration rates at 8 h (P ＞ 0.05).The expression of Vimentin,Fibronectin,MMP-2,MMP-9 and N-Cadherin in 5637 cells and T24 cells of human bladder cancer decreased significantly after adding MAP30.The E-Cadherin expression in human bladder cancer 5637 cells were decreased,but no target band was detected in human bladder cancer T24 cells.Conclusions The ribosome-inactivating protein MAP30 can effectively inhibit the migration of human bladder cancer 5637 and T24 cells by inhibiting the EMT pathway and inhibiting the expression of MMPs.
ABSTRACT
Root extract of Boerhaavia diffusa L. induced systemic resistance in tobacco against Tobacco mosaic virus. A 30 kDa protein was isolated as the active component, called BDP-30 on the basis of the molecular weight and source plant. BDP-30, a glycoprotein, was found to be temperature and protease resistant. It was basic, possessing a pI greater than 9.0. In-gel proteolytic digestion of BDP-30 generated two peptides that possessed the amino acid sequence KLYDIPPLR and KVTLPYSGNYER by LC/MS/MS. Both peptides shared absolute sequence identity with trichosanthin, a ribosome-inactivating protein from Trichosanthes kirilowii, and a 78% and 100% homology respectively with an RIP from Bryonia dioica, bryodin. Further, effort was made to look at the fate of TMV in induced resistant Nicotiana tabacum cv. Xanthi, a systemic host of the virus, at specified days after inoculation in control and treated plants. TMV coat protein (CP) was detected by immunoblot 7 days post inoculation up to 21 days in the control set, but not in treated resistant plants. TMV RNA was detected by RT-PCR using TMV-CP specific primers. Resistant tobacco did not show presence of TMV RNA up to 21 days of inoculation. This suggests that BDP-30 may be suppressing TMV replication.
ABSTRACT
Objective: To investigate the inhibitory effect of Luffin-β-targeted fusion immunotoxin, which contains endoplasmic reticulum-resident signal fragment KDEL (Lys-Asp-Glu-Leu-COOH) and the uPAcs (cleavage site of urokinase-type plasminogen activator), on NSCLC (non-small cell lung cancer) cells. Methods: Luffin-β gene was cloned by RT-PCR (reverse transcriptase-polymerase chain reaction). Fused gene Luffin-β-KDEL-uPAcs was constructed by primer extension method, and inserted in pET-32a(+) vector to form the recombinant vector pET-32a(+)/Luffin-β-KDEL-uPAcs. After pET-32a(+)/Luffin-β-KDEL-uPAcs imported into Escherichia coli, the exression of fusion protein TELKP (Trx-EK-Luffin-β-KDEL-uPAcs) was induced, and then the TELKP was purified. EK (enterokinase) was used to digest TELKP to produce the targeting fused immunotoxin LKP (Luffin-β-KDEL-uPAcs). CCK-8 (cell counting kit-8), RT-PCR and Western blotting were employed to test the cytotoxic effect of LKP cleavaged by uPA (urokinase-type plasminogen activator) for setting free immunotoxin Luffin-β on NSCLC cells in vitro. Results: The recombinant vector pET-32a(+)/Luffin-β-KDEL-uPAcs could be induced to express the fusion protein TELKP, which contained Trx tag of vector pET-32a(+) and its relative molecular mass was about 4.88×104. The immunotoxin protein LKP, which contained 290 amine acids and its relative molecular mass was about 3.1 8×1104, could be produced after the fusion protein TELKP was digested by EK. The immunotoxin Luffin-β, which possessed cytotoxic effect on tumor cells, could be released from LKP protein cleavaged by uPA in vitro, and its cytotoxic effect was dose-dependent. Conclusion: Fused gene Luffin-β-KDEL-uPAcs and its prokaryotic express vector are successfully constructed. Recombinant fusion immunotoxin LKP, whose relative molecular mass is about 3.1 8×104, is successfully prepared. The immunotoxin Luffin-β, which possesses cytotoxic effect on tumor cells, can be released from LKP protein cleavaged by uPA in vitro. Copyright © 2013 by TUMOR.