Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The 0.5 M KCl-treatment solubilizes the outer arms from sea urchin sperm axonemes. Approximately 30 percent of A-polypeptide, corresponding to dynein 1 in SDS- polyacrylamide gel, was solubilized by this treatment (as SEA-dynein 1). Electron microscopic observation indicated that the extracted axonemes lacked the outer arms in various degrees. The DEA-dynein 1 was that the extracted axonemes lacked the outer arms in various degrees. The SEA-dyenin 1 was purified and an antiserum against it was prepared in rabbits. The specificity of antiserum to dynein 1 was determined by immunoelectrophoresis and ouchterlony's double-diffusion test. The anti-dynein 1 serum inhibited ATPase activity of purified SEA-dynein 1 by 95 percent. By the indirect peroxidase-conjugated antibody method, the loci of SEA-dynein 1 within the intact, salt- extracted and mechanically disrupted axonemes were determined to be the outer arms: deposition of electron-dense materials which represents their localization was detected at the distal ends of the outer arms, in the case of intact axonemes. The 5-6 cross- bridge was hardly decorated. No decoration was seen in the salt-extracted axonemes lacking all the outer arms. In disrupted axonemes, which consist of single to several peripheral doublets, electron-dense materials were deposited only on the outer arms. Approximately 73 percent of axonemal ATPase activity sensitive to antiserum was solubilized by repeated salt-extractions. One-half of A-polypeptide (SEA-dynein 1 located at the outer arms) was contained in the pooled extracts. The extracted axonemes contained another half of A-polypeptide (SUA-dynein 1 supposed to locate at the inner arms) and retained 31 percent of axonemal ATPase activity that was almost resistant to antiserum. Solubilized SUA-dynein 1 was immunologically the same as SEA-dynein 1. This result indicates that in situ SUA-dynein 1 did not receive anti-dynein 1 antibodies, coinciding with the result obtained for salt-extracted axonemes lacking all the outer arms by the enzyme-antibody method mentioned above. These observations suggest that immunological dissimilarity in dynein 1 between outer and inner arms but do not tell us that the inner arms do not contain dynein 1.
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PMID:Immunological dissimilarity in protein component (dynein 1) between outer and inner arms within sea urchin sperm axonemes. 617 2

Alpha 1 protease inhibitor antigen was identified in the culture medium of the human ascites hepatoma cell line SK-HEP-1. Trypsin inhibitory activity and alpha 1 Pl antigen accumulated in serum-free medium concomitantly over a period of several days. Radioactive alpha 1 Pl antigen was detected in conditioned medium from cultures supplemented with 35S-L-methionine, indicating a synthesis and release of the protein. Alpha 1 Pl antigen in conditioned medium appeared to be antigenically identical to that in human plasma, and the newly synthesized (radiolabeled) antigen co-migrated with plasma, alpha 1 Pl after immunoelectrophoresis or SDS-polyacrylamide gel electrophoresis. Moreover, evidence is presented that the synthesized inhibitor exhibits functional activity, since the 35S-labeled alpha 1 Pl in conditioned medium complexes with trypsin. We conclude that SK-HEP-1 cells in culture produce functionally active alpha 1 Pl which may be identical to that in plasma.
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PMID:Functional alpha 1 protease inhibitor produced by a human hepatoma cell line. 627 80

Four fragments (F1-F4) of SEA, obtained via papain proteolysis were separated and isolated as individual components by means of the SDS-electrophoresis in polyacrilamide gel. Molecular masses of the pairs F1 + F4 and F2 + F3 are equal to the mass of the intact toxin--a fact that supposes a cleavage of polypeptide chain in two regions of "disulphide loop" in a SEA molecule. Neither fragment possesses any enterpathogenic properties. It was established, that interferonogenic and mitogenic activity of SEA is connected only with the part of molecule corresponding to F1(17,500) and F3(15,000). Two kinds of antigenic determinants in the SEA molecule were found: one was attributed to F1 and F3 fragments, the other was localised in F2 and F4. Proteolysis by trypsin led to cleavage of a small peptide from the N-terminal end of toxin molecule. Trypsinized SEA displayed all kinds of biological activity characterizing the native toxin.
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PMID:Topology of the functions in molecule of staphylococcal enterotoxin Type A. 670 70

Two IgM isotype monoclonal antibodies (McAb), 2H10 and 2H1, recognizing repetitive epitopes on Schistosoma egg-associated molecules were characterized and their specificities were identified in a two-site sandwich ELISA system. In consistent with the differences in immunological behaviour and specificity demonstrated with immunoelectrophoresis (IEP) and immunofluorescent antibody (IFA) techniques, absolutely negative reactions found in the heterologous detecting system with alternated capture and detecting McAbs of the two revealed a complete incompatibility giving evidences that epitopes on different molecules were recognized. Immunological liability of the target antigen SEA or SEA-TCA to the two McAbs were demonstrated on sodium periodate and trifluoroacetic acid treatment indicating the biochemical nature of these epitopes were glycosyolated molecules with apparently higher resistance to the oxidizing agent showing in 2H10 recognizing epitopes. By means of an ion-gradient Mono-Q FPLC system (Pharmacia), 2H10-reactive epitopes of SEA, being tested not so efficiently adsorbed by ConA-sepharose affinity column, was found successfully concentrated in the profile eluted with pH 8.0 PBS at 0.2-0.4 NaCl ionic strength. Repeated trials on SDS-PAGE and Western blotting analysis with the reactive fractions further showed a heterogeneity of molecular weight range as well as the non-transferable property of the CHO-reactive groups.
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PMID:[Analysis of the target epitopes recognized by two monoclonal antibodies directed to egg-associated fractions of Schistosoma japonicum]. 752 6

Human prostate-specific antigen (PSA), a 33 kDa kallikrein-like serine protease, occurring in the prostate, in seminal plasma and in blood, was prepared under nonreducing conditions in an enzymatically active form from seminal plasma by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by fast copper staining, electroelution from gel slices and dialysis against isotonic phosphate-buffered saline (PBS). Enzymatic activity was demonstrated for the first time directly by cleavage of semenogelin, one of the biological substrates of PSA, isolated by the same procedure, i.e. SDS-PAGE and electroelution, but from seminal vesicle fluid. The purified PSA formed SDS-stable complexes with the two major extracellular protease inhibitors in blood, alpha 1-antichymotrypsin (alpha 1-ACH) and alpha 2-macroglobulin (alpha 2-M). PSA isolated under reducing conditions was enzymatically inactive and could not bind to the protease inhibitors alpha 1-ACH and alpha 2-M.
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PMID:Biological activity of prostate-specific antigen isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electroelution. 758 64

We have reported that overexpression of Neu leads to heregulin-stimulated neurite outgrowth and the tyrosine-phosphorylation of Neu and other cellular proteins in PC12 cells. Considering that Neu/ErbB2 alone is not able to functionally couple to heregulin, we looked for the possible involvement of ErbB3 in these neurite outgrowth and tyrosine phosphorylation responses. We found that heregulin stimulates the tyrosine phosphorylation of endogenous ErbB3 protein in PC12 cells and that this phosphorylation, like that of Neu, is greatly enhanced in cells that overexpress Neu. Furthermore, overexpression of ErbB3 in PC12 cells led to heregulin-stimulated neurite extension. In addition to becoming tyrosine-phosphorylated, Neu/ErbB2 and ErbB3 associate with each other, and each associates with the 85-kDa regulatory subunit (p85) of phosphatidylinositol 3-kinase in a heregulin-dependent manner. Thus, Neu/ErbB2 and ErbB3 appear to cooperate to mediate the heregulin signal in PC12 cells. Like heregulin, epidermal growth factor (EGF) also stimulate the tyrosine phosphorylation of both Neu and ErbB3. However, there are clear differences between the EGF- and heregulin-stimulated phosphorylations of ErbB3. In the heregulin response, two tyrosine-phosphorylated forms of ErbB3 are detected. Of these, only the more quickly migrating form (on SDS-polyacrylamide gel electrophoresis) is found to be associated with Neu, whereas the other, more slowly migrating form is uniquely capable of forming stable complexes with p85. In the EGF response, at least two tyrosine-phosphorylated forms of ErbB3 are detected, but these phosphoproteins have distinctly lower apparent molecular weights compared with the heregulin-stimulated ErbB3 phosphoproteins and do not complex with p85. Thus the formation of a stable ErbB3-p85 complex in PC12 cells is a unique outcome of heregulin signaling that correlates with the differences in cell morphology induced by the activated EGF receptor and the Neu tyrosine kinase.
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PMID:Heregulin-stimulated signaling in rat pheochromocytoma cells. Evidence for ErbB3 interactions with Neu/ErbB2 and p85. 764 63

The primary biological effect of the estrogen estradiol-17 beta (17 beta E2) on bone is to decrease bone resorption. However, whether 17 beta E2 affects osteoclast differentiation or function directly or through its action on osteoblasts is unclear. To investigate this question we examined the human preosteoclastic cell line FLG 29.1 for evidence of functional estrogen receptors (ERs). Southern blotting of reverse transcription-PCR amplification products with a 32P-labeled cDNA probe for the human ER mRNA demonstrated that FLG 29.1 cells express ER mRNA. Binding of [3H]17 beta E2 to nuclear ERs was steroid specific with approximately 400 saturable, high affinity (Kd approximately 1 nM) binding sites per cell nucleus. Nuclear ERs covalently labeled with [3H]tamoxifen aziridine showed an apparent molecular weight of 65,000 by SDS/PAGE and Western blotting with the D75 monoclonal antibody to human ER. Pretreatment of cells with 0.1, 1.0, or 10 nM 17 beta E2 induced a dose- and time-dependent specific binding of progesterone to FGL 29.1 cells, and stimulation of the cells with 10 nM and 100 nM 17 beta E2 significantly (P < 0.05) reduced cell proliferation. Transcriptional activity of the ER gene was detected by transient transfection of cells with the pERE-BLCAT plasmid containing the estrogen response element for the vitellogenin A2 gene and the bacterial chloramphenicol acetyltransferase reporter gene. Treatment of FLG 29.1 cells with 10 nM 17 beta E2 increased chloroamphenicol acetyltransferase expression from 5- to 29-fold compared to controls. These observations suggest a potential role for estrogen in osteoclastogenesis.
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PMID:Functional estrogen receptors in a human preosteoclastic cell line. 770 3

In the present work we have investigated activation of phosphorylation of plasma membrane insulin-like growth factor-I receptors (IGF-IR) by diethylstilbestrol (DES). Insulin-like growth factor-I (IGF-I) stimulated the activity of membrane protein tyrosine kinase(s) (PTK) in both normal and DES-treated hamster kidneys. The level of IGF-I-stimulated PTK(s) almost doubled after 15 days of DES treatment. Autophosphorylation experiments revealed that phosphorylation of a 95 kDa band (presumably the beta subunit of IGF-IR) was 2-fold higher in the membranes of kidney from DES-treated animals compared with controls. To understand the mechanism of activation of IGF-I-dependent PTK by DES, we investigated the relationship between the binding capacity of IGF-I to membrane proteins and the level of IGF-IR. The binding of [125I]IGF-I to membranes from the DES-treated group was 30% higher than that of age-matched normal kidney (P < 0.001). Scatchard analysis of the binding data for both normal and DES-treated hamster kidney revealed a single class binding site for IGF-I with a dissociation constant (Kd) of 4.1 and 4.6 nM and a maximum binding capacity (Bmax) of 1786 and 2086 fmol/200 micrograms protein respectively. Therefore, the difference observed in [125I]IGF-I binding between DES-treated and normal kidney membranes may be partially due to an increase in the number of IGF-I binding sites, with no change in the affinity of the receptors for IGF-I. An enhanced level of IGF-IRs in membranes from DES-treated animals was visualized by autoradiography following affinity labeling of membrane proteins subjected to SDS-PAGE. Under reducing conditions a molecular band of 132 kDa was evident. The 132 kDa band represents the alpha-subunit of IGF-IRs. Northern blot analyses revealed that DES treatment increased the level of IGF-IR mRNA 2-fold compared with that of controls. These findings suggest that an enhanced level of IGF-IR coupled with qualitative changes may be responsible for the activation of IGF-I-dependent PTK on DES exposure. Whether the stimulation of IGF-IR phosphorylation by exposure to a carcinogenic dose of DES may be a factor in the induction of renal cancer in Syrian hamsters is not clear.
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PMID:Activation of phosphorylation of plasma membrane insulin-like growth factor-I receptors in the kidney of Syrian hamsters by diethylstilbestrol. 778 52

Major diagnostic proteins of 31/32 kDa were purified from soluble adult worm homogenate of Schistosoma japonicum using AcA ultragel chromatography and additional treatments. The purity of the isolated antigenic proteins (Sj 31/32) was demonstrated by SDS-PAGE and Western blotting technique. The antigens showed a negative reaction when stained by PAS and remained active after treatment with sodium periodate, moving toward anode in the electrophoretic field. These purified antigens were used for detecting the specific antibody in sera from patients with schistosomiasis japonica by ELISA and IHA. As compared with SEA, Sj 31/32 kDa was found as sensitive as but much more specific than SEA in immunodiagnosis.
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PMID:Purification of 31/32 kDa proteins of adult Schistosoma japonicum as antigens (Sj 31/32) for ELISA and IHA. 778 90

Vascular endothelial cell growth factor (VEGF), an endothelial cell-specific mitogen that plays an important role in angiogenesis, promotes the tyrosine phosphorylation of at least 11 proteins in bovine aortic endothelial cells (BAEC). Proteins immunoprecipitated from lysates of control- and VEGF-stimulated BAEC with antisera to phospholipase C-gamma (PLC-gamma) were fractionated by SDS-polyacrylamide gel electrophoresis and transferred to Immobilon-P. Evaluation of the Western blots with antisera to phosphotyrosine demonstrated that PLC-gamma and two proteins (100 and 85 kDa) that associate with PLC-gamma were phosphorylated in response to VEGF. By using antisera specific to other mediators of signal transduction that contain SH2 domains for immunoprecipitation, it was demonstrated that VEGF promotes phosphorylation of phosphatidylinositol 3-kinase, Ras GTPase activating protein (GAP), and the oncogenic adaptor protein NcK. Proteins of M(r) consistent with the VEGF receptors Flt-1 and Flk-1/KDR were also tyrosine phosphorylated in stimulated cells. Tyrosine-phosphorylated Nck, PLC-gamma, and two GAP-associated proteins, p190 and p62, were in GAP immunoprecipitates of VEGF-stimulated BAEC, and tyrosine-phosphorylated NcK was in phosphatidylinositol 3-kinase immunoprecipitates. These observations suggest that VEGF promotes formation of multimeric aggregates of VEGF receptors with proteins that contain SH2 domains and activate various signaling pathways. VEGF-promoted proliferation of endothelial cells and tyrosine phosphorylation of SH2 domain containing signaling molecules were inhibited by the tyrosine kinase inhibitor genistein.
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PMID:Vascular endothelial cell growth factor promotes tyrosine phosphorylation of mediators of signal transduction that contain SH2 domains. Association with endothelial cell proliferation. 789 17


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