Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.108 (TAT)
 2,389 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Vascular heparan sulfate proteoglycan (vHSPG) is an important functional component of the microvasculature. Previous studies have demonstrated autoimmunity to vHSPG in systemic lupus erythematosus (SLE). In the current studies, we further investigated the immunospecificity of anti-vHSPG antibodies in SLE sera by enzyme-linked immunoassay (ELISA). In direct binding assays, SLE sera contained IgG antibodies reactive with native vHSPG and with heparan sulfate (HS) glycosaminoglycan in significantly higher titers than controls. Employing purified SLE IgG in liquid-phase competitive immunoinhibition ELISAs, SLE IgG anti-HS antibodies cross-reacted with heparin and DNA, but not with other glycosaminoglycans or anionic phospholipid antigens. Immunochemical studies demonstrated that the immunodominant site on HS recognized by SLE IgG contained 2-O-sulfated uronic acid. Removal of N-sulfated and 6-O-sulfated residues primarily on N-acetyl-glucosamine had no effect on antigenicity, further demonstrating that nonspecific charge interactions which are the result of sulfation do not solely account for the antigenicity of HS. SLE IgG from patients with active SLE was further affinity purified on DNA-cellulose and HS-Sepharose columns for immunospecificity studies. After affinity purification of both anti-DNA and anti-HS antibodies, significant enhancement of direct binding reactivity with HS was noted. In addition, anti-DNA and anti-HS IgG antibody reacted with the cell surface of endothelial cells by a cellular ELISA (CELISA). Immunoinhibition studies of CELISA reactivity confirmed that affinity-purified SLE IgG anti-DNA anti-HS antibody were reactive with endothelial cell surface HS antigens. Furthermore, SLE IgG anti-DNA antibody reactivity with endothelial cells was not reduced by DNase treatment of the cells, but was significantly reduced by heparitinase digestion. Since HS plays an important role in the maintenance of normal anticoagulation on the endothelial cell surface by binding antithrombin III, we investigated the inhibition of heparin-accelerated thrombin-antithrombin III complex formation by SLE IgG. Purified IgG from patients with active SLE, but not from normal controls, inhibited heparin-accelerated formation of TAT complexes. These studies demonstrate the presence of IgG autoantibodies to HS in patients with SLE. Anti-HS antibodies recognize an antigenic site also present in heparin, but not other glycosaminoglycans, bind to the endothelial cell surface, and inhibit the formation of TAT complexes. SLE IgG anti-HS antibodies recognize a sulfated uronic acid epitope containing 2-O-sulfate which is important in certain functions of HS, including antithrombin III binding. Thus, anti-HS antibodies may promote a procoagulant state at the endothelial cell surface.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Autoantibodies to vascular heparan sulfate proteoglycan in systemic lupus erythematosus react with endothelial cells and inhibit the formation of thrombin-antithrombin III complexes. 829 26

Cancer suicide gene therapy based on herpes simplex virus type I thymidine kinase (HSV-TK) and ganciclovir (GCV) suffers from the lack of efficacy in clinical use, which is mostly due to low gene-transfer efficiency and absence of bystander effect in tumors. We have previously demonstrated the enhancement of GCV cytotoxicity by fusing the HSV-TK with the cell penetrating peptide from HIV-1 transactivator protein transduction domain (TAT PTD). Despite the earlier promising results, we found that the triple fusion protein HIV-1 transactivator protein transduction domain-thymidine kinase suicide gene-green fluorescent protein marker gene (TAT-TK-GFP) increased GCV cytotoxicity only in 3/12 of different human tumor cell lines. Extended GCV exposure enhanced the cytotoxic effect of HSV-TK/GCV gene therapy, but the difference between TK-GFP and TAT-TK-GFP was not statistically significant. The modest improvement on cell killing mediated by TAT PTD in Chinese hamster ovary cells appeared to be associated with cell-surface heparan sulfate proteoglycan (HSPG) composition. However, TAT-mediated increased cell death did not correlate with the density of cell-surface HSPG expression in different tumor cell lines. In conclusion, although some degree of enhancement by TAT was shown in certain tumor cells in vitro, it is unlikely that TAT peptide linked to a suicide protein could be a useful booster of in vivo gene therapy trials.
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PMID:Characterization of HIV-1 TAT peptide as an enhancer of HSV-TK/GCV cancer gene therapy. 1830 53

Biomolecules such as proteins, DNA, and RNA are macromolecules and can not cross the cell membrane. However, cell-penetrating peptide (CPP) has been shown to deliver therapeutic biomolecules successfully into cells. The various and widely used CPPs including TAT, VP22, and Antp are mostly non-human originated CPPs, and are limited by their potential toxicity and immunogenicity. We report here on a newly identified novel cell-penetrating sequence (LPIN; RRKRRRRRK) from the nuclear localization sequence (NLS) of human nuclear phosphatase, LPIN3. LPIN-EGFP recombinant protein was concentration- and time-dependently delivered into cells and localized to the nucleus as well as the cytoplasm. It penetrated the cell membrane by lipid raft-mediated endocytosis by binding to heparan sulfate proteoglycan. LPIN-EGFP was successfully delivered into primary mouse splenocytes in vitro and it could be delivered into various tissues including liver, kidney, and intestine in mice after intra-peritoneal injection. This research suggests that LPIN-CPP could be used in a drug delivery system to deliver therapeutic biomolecules including peptides, proteins, DNA, and RNA and without the limitations of non-human originated CPPs such as TAT-CPP.
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PMID:Identification of a novel cell-penetrating peptide from human phosphatidate phosphatase LPIN3. 2326 58

We have previously reported that cationic poly-arginine and arginine-rich cell-penetrating peptides display high-level neuroprotection and reduce calcium influx following in vitro excitotoxicity, as well as reduce brain injury in animal stroke models. Using the neuroprotective peptides poly-arginine R12 (R12) and the NR2B9c peptide fused to the arginine-rich carrier peptide TAT (TAT-NR2B9c; also known as NA-1), we investigated the mechanisms whereby poly-arginine and arginine-rich peptides reduce glutamate-induced excitotoxic calcium influx. Using cell surface biotin protein labeling and western blot analysis, we demonstrated that R12 and TAT-NR2B9c significantly reduced cortical neuronal cell surface expression of the NMDA receptor subunit NR2B. Chemical endocytic inhibitors used individually or in combination prior to glutamate excitotoxicity did not significantly affect R12 peptide neuroprotective efficacy. Similarly, pretreatment of neurons with enzymes to degrade anionic cell surface proteoglycans, heparan sulfate proteoglycan (HSPG), and chondroitin sulfate proteoglycan (CSPG), as well as sialic acid residues, did not significantly affect peptide neuroprotective efficacy. While the exact mechanisms responsible for R12 peptide-mediated NMDA receptor NR2B subunit cell surface downregulation were not identified, an endocytic process could not be ruled out. The study supports our hypothesis that arginine-rich peptides reduce excitotoxic calcium influx by reducing the levels of cell surface ion channels.
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PMID:The Neuroprotective Peptide Poly-Arginine-12 (R12) Reduces Cell Surface Levels of NMDA NR2B Receptor Subunit in Cortical Neurons; Investigation into the Involvement of Endocytic Mechanisms. 2786 26