[Effect of cultivation in zinc media on the growth of and the degree of zinc in organic form in transgenic metallothionein mushroom].

Metallothioneins (MTs) are a family of low molecular weight, cysteine-rich and metal-binding functional proteins. Transgenic MT mushroom can be used as functional food additives, but its zinc-enriching ability has not been studied systemically until now. The zinc contents in mycelia of transgenic MT mushroom (Pleurotus ostreatus) and wild type mushroom mycelia cultivated in different zinc concentration media were analyzed by ICP-OES.

[Distribution of Pb and Zn in transgenic metallothionein tobacco].

Transgenic metallothionein (MT) plant can clear the heavy metals from soil and environment, but the distribution of metals in plants has not been studied systematically. The Pb and Zn contents in different parts of transgenic MT tobacco plant of sixth generation and traditional plant (same culture variety as control) were analyzed. The Pb and Zn contents in total transgenic plant were 21.

A potential thrombus diagnosis reagent based on P-selectin monoclonal antibody SZ-51 light chain.

Introduction: P-selectin is a well characterized platelet adhesion molecule that can shift from the secretory granules to the surface of activated platelets, which makes it a potential target in thrombus diagnosis and therapy. SZ-51 is a monoclonal antibody against P-selectin.

Materials And Methods: To build a potential thrombus diagnosis reagent, we expressed the light chain of SZ-51 (SZ-LC) in P.

[Decontamination of heavy metals in wastewater by transgenic Synechococcus sp. PCC 7002 with mouse metallothionein-I gene].

The growth characteristics and the decontamination of heavy metals in analogous heavy metals wastewater by transgenic Synechococcus sp. PCC 7002 with mouse metallothionein-I gene were studied. The results show that transgenic Synechococcus sp.

Improving zinc content and antioxidant activity in transgenic tomato plants with expression of mouse metallothionein-I by mt-I gene.

Metallothioneins (MTs), as a family of low-molecular-weight, cysteine-rich, and metal-binding proteins, show potential for utilization in functional food. Tomato plants were transformed with gene constructs that contained mt-I encoding the mouse MT-I, similar in sense orientation with the constitutively active double 35S promoter from cauliflower mosaic virus. Three independent transformants, which had copies of the gene in their genomes, were obtained.

Survival of human metallothionein-2 transplastomic Chlamydomonas reinhardtii to ultraviolet B exposure.

Solar ultraviolet (UV) radiation has a great influence on green organisms, especially plankton like Chlamydomonas. A human metallothionein-2 gene, which is generally considered to have an anti-radiation function by its coding product, was transferred into the chloroplast genome of Chlamydomonas reinhardtii. To dynamically measure the UV effects on Chlamydomonas cells grown in liquid tris-acetate-phosphate medium, a new method was developed based on the relationship between the chlorophyll content of an algal culture and its absorbance at 570 nm after the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.

Studies on the epitope of neuronal growth inhibitory factor (GIF) with using of the specific antibody.

Human neuronal growth inhibitory factor (GIF), a metalloprotein classified as metallothionein-3, is specifically expressed in mammal central nervous system (CNS). In these Studies the specific antibody to human GIF was prepared and used to search the epitope of human GIF by enzyme-linked immunosorbent assay (ELISA) and sequence comparison. The result of ELISA showed the epitope of human GIF may locate on a octapeptide (EAAEAEAE) in the alpha-domain of human GIF, and the result of nerve cell culture indicated that the biological activity of GIF may be affected by the specific antibody.

Proteomic analysis of corticobasal degeneration: a case study of corticobasal degeneration at the proteome level.

Corticobasal degeneration (CBD) is an adult-onset progressive neurodegenerative disorder. However, its pathogenic mechanisms underlying this disorder remain poorly understood. The authors examined changes of proteome profiles between three nondemented comparison brains and a CBD brain based on two-dimensional gel electrophoresis.

Characterization of expression and stability of recombinant cystein-rich protein human MT1A from yeast.

Metallothionein (MT) is the protein that has been shown to bind heavy metals, scavenge free radicals, protect DNA from radiation damage, and alleviate disease symptoms. However, only very limited success has been achieved in expression and production of active recombinant metallothionein. In this study, human metallothionein 1A (hMT1A) was transformed into yeast Pichia pastoris for expression with secretion of the protein into the medium.

Effect of metallothionein on cell viability and its interactions with cadmium and zinc in HEK293 cells.

Metallothioneins (MTs) are thought to participate in a wide variety of physiological roles, but the mechanisms involved are still unclear. The study was designed to examine the possible factors related to these mechanisms. Methods, including transfection, MTT assay and flow cytometry, were used to investigate the effect of MTs on cell viability and their interactions with cadmium and zinc in HEK293 cells.

[Affection of metallothionein-3 to dUTPase's accommodating cellular toxicity of dUTP].

Metallothionein-3 (MT-3), renamed as growth inhibitory factor (GIF), is a brain specific member of the metallothionein family. Human dUTPase is a recently found protein in brain that can interact with hMT-3. They have the growth inhibitory activity on neuron cell by interaction.

High-level expression of human TFF3 in Escherichia coli.

A strategy for expression and purification of recombinant N-terminal human trefoil factor family-domain peptide 3 (hTFF3) in Escherichia coli was established. The gene of hTFF3 was synthesized to substitute the low-usage condons with corresponding high-usage synonymous condons. At the same time, the signal peptide of DsbC was added to the N-terminus of the hTFF3 gene.

Expression of an antibody-targeted plasminogen activator in Escherichia coli using chemically induced stress responses.

A recombinant gene coding for an antibody-targeted urokinase-type plasminogen activator was constructed for the purpose of enhancing the thrombolytic specificity of urokinase. The recombinant gene was cloned into prokaryotic expression vector pTrcHisA, and transformed into Escherichia coli strain Rosetta (DE3). Less than 4 mg of the desired protein l(-1) could be obtained in the form of inclusion bodies.

[Molecular cloning and identification and biological activity of GIF-interacting protein(s)].

In order to examine the elusive functional mechanism of GIF (Neuronal growth inhibitory factor, GIF) and elucidate the possible relationship between GIF and Alzheimer's disease, we constructed bait1 plasmid (pHyblex-GIF) by cloning GIF cDNA directly in frame with plasmid pHyblex, and used the yeast two-hybrid system to screen Alzheimer's disease human brain cDNA library and found the GIF-interacting proteins. The final results from coimmunoprecipitation and western blotting experiments confirmed that interacting proteins specifically binds to GIF. After sequencing the nucleotide of the putative positive plasmids and searching for homologues, we found that one of these is the part of human nuclear dUTPase protein sequence.

Functional properties of a recombinant chimeric plasminogen activator with platelet-targeted fibrinolytic and anticoagulant potential.

The construction, purification, and characterization of dscuPA33khC, a bifunctional protein designed for thrombosis treatment is described. The chimera was designed to consist of a decorsin (platelet aggregation inhibitor), a low molecular mass (33kDa) single-chain urokinase (scuPA-33k), and a thrombin inhibitory domain. We have successfully produced this recombinant protein in the Escherichia coli expression system, in which the target protein exists in the form of inclusion bodies.

Enhanced expression of an antibody-targeted plasminogen activator in Escherichia coli by fusion to decorsin.

To improve the thrombolytic specificity of plasminogen activators, an antibody-targeted plasminogen activator was constructed consisting of a single-chain variable fragment of a monoclonal antibody SZ-51 raised specifically against human P-selectins on activated platelets and a low molecular weight single-chain urokinase. After fusion to the 3' end of the gene coding for decorsin, originally isolated from the leech Macrobdella decora, expression of the antibody-targeted plasminogen activator gene in E. coli strain Rosetta (DE3) pLysS was greatly enhanced.

Bacterial expression and purification of biologically active human TFF3.

A glutathione S-transferase (GST) fusion protein expression system for the production and purification of recombinant human trefoil factor family-domain peptide 3 (hTFF3) was established. The hTFF3 gene, prepared by PCR, was cloned into a pBluescript KS(+) plasmid, and inserted into a pGEX-4T-1 GST fusion vector. The GST-hTFF3 fusion protein was expressed in Escherichia coli, and hTFF3 was purified with Glutathione Sepharose 4B affinity chromatography, yielding about 3-4 mg of pure hTFF3 in one liter of culture broth.

Expression and characterization of ARSP1 from Eisenia fetida.

To study the characterization of a protease ARSP1 (apoptosis-related serine protease) of Eisenia fetida, a recombinant ARSP1 was constructed. ARSP1 was produced in E. coli BL21-CodonPlus (DE3)-RIL after IPTG induction and exited in inclusion body.

Attenuation of zinc-induced neuronal death by the interaction of growth inhibitory factor with Rab3A in rat hippocampal neurons.

We examined the protective effect of growth inhibitory factor (GIF) against zinc-induced neuronal death in rat hippocampal neurons. In an in vitro cell culture system, 300 microM Zn(2+) readily induced death of hippocampal neuronal cells, which was characterized by massive necrosis and a minor degree of apoptosis. Neither the addition of recombinant GIF nor Rab3A alone could rescue these cells from death.

Differential display proteome analysis of PC-12 cells transiently transfected with metallothionein-3 gene.

Metallothionein-3 (MT-3), also known as growth inhibitory factor, possesses several unique properties other than the common features of metallothionein family. To investigate the mechanisms underlying its multifaceted roles in the central nervous system, we employed differential display proteomics techniques to study holistic protein changes of PC-12 cells induced by transient transfection of MT-3. Ten significantly and reproducibly changed proteins were identified and their functional implications are discussed in some detail.

[Effect of recombinant human trefoil factor on protection of gastric mucosa and healing of chronic gastric ulcer in rats].

Objective: To study the effect of recombinant human intestinal trefoil factor ( rhITF) on the healing of rat chronic gastric ulcer, protect gastric mucosal and mechanisms are involved.

Methods: (1) Acute gastric mucosal injury was induced by ethanol, stress, aspirin and pylorusl ligation. The injury index,MDA, GMBL,hexosamine (Hex) and acid output were measure.

Differential protein expression induced by transient transfection of metallothionein-3 gene in SH-SY5Y neuroblastoma cell line.

Metallothionein-3(MT-3), also known as growth inhibitory factor (GIF), is predominantly expressed in central nervous system (CNS). It belongs to the family of metallothionein(MT) but has several unique properties that are not shared by other family members such as MT-1/MT-2. In the past few years, MT-3 had been postulated to be a multipurpose protein which could play important neuromodulatory and neuroprotective roles in CNS besides the common roles of MTs.

Proteomic comparison between human young and old brains by two-dimensional gel electrophoresis and identification of proteins.

To investigate molecular mechanisms of human brain aging, brain proteins were isolated from postmortem human young and old brains and profiled by two-dimensional gel electrophoresis (2-DE). With the help of special software, five down-regulated protein spots in two-dimensional gel electrophoresis gels of old brains were found compared with young brains, four of which was identified as a protein similar to peroxiredoxin 2 (accession-numbered as gi | 13631440), two of stathmin (phosphoprotein p19) and apolipoprotein A-I precursor (apo-AI) by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Eight common proteins, whose expressions were not altered between young and old brains, were also identified.

Rheb is a direct target of the tuberous sclerosis tumour suppressor proteins.

Mutations in the TSC1 or TSC2 genes cause tuberous sclerosis, a benign tumour syndrome in humans. Tsc2 possesses a domain that shares homology with the GTPase-activating protein (GAP) domain of Rap1-GAP, suggesting that a GTPase might be the physiological target of Tsc2. Here we show that the small GTPase Rheb (Ras homologue enriched in brain) is a direct target of Tsc2 GAP activity both in vivo and in vitro.