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)

Previous studies identified three COOH-terminal residues in staphylococcal enterotoxin E (SEE; Asp200, Pro206, and Asp207) that in part mediate TCR V beta recognition. We have identified an additional three residues near the NH2-terminus of SEE (Arg20, Asn21, and Ser24 that are needed in conjunction with these COOH-terminal residues to fully restore native levels of V beta-specific T cell proliferation. A staphylococcal enterotoxin A SEA-SEE hybrid molecule containing the NH2-terminal V beta determinants of SEE to activate alone exhibited V beta specificities of both SEA and SEE, indicating that these residues of SEE independently contribute to V beta recognition and do not obscure the native V beta determinants of SEA. These findings suggest that the ability of SEE to activate certain V beta-specific T cell subsets may result from multiple interactions with a single TCR beta-chain or perhaps by cross-linking two TCR. High affinity binding to HLA-DR1, a property of native SEA, was not altered in the SEA-SEE hybrid enterotoxins containing amino acid substitutions in regions 20 to 24 and 200 to 207, indicating that residues comprising the V beta determinants of SEE are separate from residues that contribute to HLA-DR1 binding affinity. Computer models of the predicted structure of SEE revealed that the V beta determinants of SEE are located on two adjacent solvent-exposed loops. Thus, the residues of SEE that mediate V beta recognition may coalesce to form a TCR binding site with specificities for multiple TCR beta-chains.
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PMID:Residues near the amino and carboxyl termini of staphylococcal enterotoxin E independently mediate TCR V beta-specific interactions. 869 Sep 7

The RIDASCREEN SET kit (R-Biopharm GmbH, Darmstadt, Germany), a commercial staphylococcal enterotoxin (SE) immunoassay kit, utilizes monovalent capture antibodies against SE types A to E (SEA to SEE); therefore, it simultaneously detects and identifies the enterotoxin types. A collaborative study was conducted to ascertain whether the specificity, sensitivity, repeatability and reproducibility of the kits would meet food safety criteria. Twelve Canadian laboratories participated in this study to analyze various foods to which 1.0 to 2.0 ng of SE/g had been added and negative control samples. The results indicate that the sensitivity and specificity of the kit were excellent; all collaborators were able to detect the minimum toxin levels of 1.0 ng SEA/g in ham and cheese, 1.0 ng SEB/g in salami and turkey, and 2.0 ng SED/g in other samples without any false-negative results. With regard to negative control samples, all analysts obtained correct results except for one analyst who recorded weak false-positive results with several foods detecting SEC or SEA. The overall rate of false-positive results was 0.7% for 600 triplicate assays. In addition, it was confirmed that the RIDASCREEN kit did not yield false-positive results with mussels in contrast to some other EIA kits. Overall repeatability and reproducibility of the kit were in the range of 11.69-42.57% and 17.25-68.05%, respectively.
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PMID:A collaborative study on the detection of staphylococcal enterotoxins in foods by an enzyme immunoassay kit (RIDASCREEN). 879 29

Adoptive transfer of tumor-sensitized T lymphocytes has demonstrated therapeutic efficacy in animal tumor models and in some patients with melanoma and renal cell cancers. In experimental settings, T lymphocytes derived from lymph nodes (LNs) draining progressively growing tumors can be activated ex vivo to generate tumor-reactive lymphocytes with therapeutic efficacy. Despite the theoretical concern regarding inaccessibility of the central nervous system to systemically transferred T cells, our recent experiments demonstrated that anti-CD3-activated tumor-draining LN cells are capable of mediating the regression of established intracerebral tumors. In this study, several staphylococcal enterotoxins (SEs), including SEA, SEC2, and SEE, and exfoliating toxin, known to be superantigens, were tested for their ability to stimulate tumor-draining LN cells to acquire antitumor reactivity for the treatment of intracerebral tumors. SEs bind to the MHC class II molecule and provide an activating signal to T cells bearing particular T-cell receptor Vbeta chains. Tumor-draining LN cells activated with SEs demonstrated selective Vbeta T-cell expansion. In adoptive immunotherapy of intracranial (IC) tumors, SEA-and SEC2-activated cells had the highest efficacy, whereas SEE-activated cells were not therapeutic. Despite the antigen independence of SE activation, the T cells retained immunological specificity for the tumor, which provided the initial in vivo sensitization of the LN. During the ex vivo stimulation with superantigens, both CD4+ and CD8+ T cells proliferated, and both subsets were required to mediate regression of IC tumors. In contrast to the adoptive immunotherapy of visceral tumors, the systemic administration of exogenous interleukin 2 failed to support the antitumor reactivity in mice depleted of CD4 cells, and, in fact, it inhibited the therapeutic efficacy. Furthermore, mice cured of intracerebral tumors by the adoptive transfer of T cells were resistant to an IC tumor rechallenge. However, in contrast to the immunological specificity demonstrated during the primary adoptive transfer, cured mice were able to reject challenge with several immunologically distinct fibrosarcomas but not a melanoma. These results indicate that superantigen-activated LN cells can circulate to and interact with intracerebral tumors mediating tumor regression in an immunologically specific manner. Although conditions that optimize the treatment of intracerebral tumors appear to be different from those for visceral tumors, analysis of T-cell receptor Vbeta expression among cells activated with several superantigens does not reflect a preferential usage of Vbeta gene segments in the immune response to autochthonous tumors.
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PMID:Treatment of intracranial tumors by systemic transfer of superantigen-activated tumor-draining lymph node T cells. 884 Sep 87

The superantigens staphylococcal enterotoxin A and E (SEA and SEE) both contact major histocompatibility complex (MHC) class II molecules on two sites located on the alpha and beta chains. We have investigated the role of the T cell receptor (TCR) alpha chain in the modulation of the various topologies of TCR/SEA (or SEE)/class II complexes. For this purpose, we have used three mouse V beta20 T cell lines expressing different V alpha domains and two T cell hybridomas expressing mouse V beta1 or V beta11 segments. The response of these T cells to SEA and SEE was studied in the context of presentation by wild-type human MHC class II molecules; or by mutants on MHC, in each of the two superantigen binding sites (position alpha39K and beta81H) to which the superantigens can still bind but with an altered conformation. Although V beta20 T cell lines are efficiently stimulated using SEA and SEE presented by wild-type HLA-DR1 molecules, our results show that the nature of the TCR V alpha domain can affect differently the recognition of the toxins bound to mutant class II molecules. This suggests that various functional topologies exist for both SEA and SEE/class II complexes and that the T cell response to each of these complexes can be modulated by the V alpha domain of the TCR. Interestingly, the recognition of SEA and SEE is achieved in different fashions by a given V beta20 T cell line.
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PMID:V alpha domain modulates the multiple topologies of mouse T cell receptor V beta20/staphylococcal enterotoxins A and E complexes. 902 3

Hypersensitivity pneumonitis (HP) and sarcoidosis are interstitial lung disorders (ILD) characterized by a lymphocytic alveolitis that, in the active phase of the disease, is sustained by different T-cell subsets, i.e., CD8+ cells in HP and CD4+ lymphocytes in sarcoid patients. To address the question of whether a bias in T-cell selection occurs in the lung of patients with HP and sarcoidosis, we analyzed the T-cell receptor beta chain variable region (TCR-Vbeta) repertoire by flow cytometry and polymerase chain reaction (PCR) analyses in blood and lung lymphocytes of 14 HP and 25 sarcoid patients. To verify whether these cells can be activated in vitro through the TCR, blood and lung lymphocytes were also assessed for their responsiveness to different superantigenic stimuli represented by staphylococcal enterotoxins, including SEA, SEB, SEC1, SEC2, SED, and SEE. Flow cytometry and PCR analyses demonstrated an overexpression of cells bearing Vbeta2, Vbeta3, Vbeta5, Vbeta6, and Vbeta8 gene segments in the lung of HP patients as compared with the peripheral blood. In sarcoid patients cells bearing Vbeta2, Vbeta5, and Vbeta6 gene segments in the lung of HP patients as compared with the peripheral blood. In sarcoid patients cells bearing Vbeta2, Vbeta5, and Vbeta6 gene segments were overrepresented in the lung rather than in the blood. Both in HP and sarcoid patients almost all T cells bearing the dominant Vbeta segment belonged to the T-cell subset that sustains the alveolitis, i.e., CD8 in HP patients and CD4 in sarcoid subjects. Follow-up studies demonstrated that the recovery of the alveolitis was characterized by the disappearance of cells bearing a limited T-cell repertoire. Interestingly, T-lymphocyte response to different superantigens demonstrated that the proliferation elicited by different staphylococcal toxins was more pronounced in the lung than in the blood. Taken together, our findings indicate a compartmentalization of cells bearing discrete Vbeta gene products in the pulmonary microenvironment and suggest that the expansion of specific Vbeta region subsets occurring in the lung might result from triggering by a specific antigen. In fact, the removal from exposure in HP patients or specific treatment in sarcoidosis resulted in the decrease of the overrepresented cell population accounting for the lymphocytic alveolitis.
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PMID:Selection of T lymphocytes bearing limited TCR-Vbeta regions in the lung of hypersensitivity pneumonitis and sarcoidosis. 903 99

In this report, we show that despite an overall amino acid residue identity of more than 80% between the staphylococcal enterotoxins (SE) A and E, these proteins markedly differ in their absolute requirement for the MHC class II during T cell activation. The superantigens were produced as C215Fab-SE fusion proteins and analyzed for their ability to activate T cells in a MHC class II-independent manner, using C215 Ag expressing cell lines as pseudo super-APCs. C215Fab-SEA, but not C215Fab-SEE, induced T cell cytotoxicity and proliferation in these MHC class II-independent systems. Introduction of a region from SEA, comprising amino acids 20-27, to SEE transferred the ability to engage T cells in the absence of MHC class II. Analysis of the Vbeta specificity of the chimeric SEA/SEE molecules and a panel of SEA mutants demonstrated that the site for TCR interaction covers the edge surrounding the shallow cavity on top of the SEA molecule.
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PMID:Functional characterization of the interaction between the superantigen staphylococcal enterotoxin A and the TCR. 912 86

The staphylococcal enterotoxins, SEA and SEE, bind one zinc atom per molecule of protein. The presence of this metal atom enhances the binding of the toxins to MHC class II molecules, presumably through an interaction with histidine 81 of the beta chain. L cell transfectants expressing HLA-DR1 and HLA-DR7 molecules, with mutations in either the alpha1 or beta1 domains, were tested for their ability to bind SEA and present it to T cells. Cells expressing DR1 molecules with alanine at positions 77, 78, 80, 83, 84 and 85, or serine at position 79 could all bind SEA and present it to either polyclonal or monoclonal T cells. Most point mutations within the alpha-helical portion of the DR7 beta chain had no effect on binding and presentation. However, substitution of histidine 81 with alanine, glutamate, or aspartate, abrogated SEA binding as well as T cell stimulation by the superantigen. This effect was also observed when the non-polymorphic aspartate, at position 76 was changed to alanine. Mutation of the asparagine at position 82 had an intermediate effect. Point mutations of the DR alpha chain had little effect on binding of SEA as determined by a flow cytometric assay. However, mutation of lysine at position 39 of the alpha chain and, to a lesser extent methionine at position 36, markedly decreased the ability of SEA to stimulate toxin-responsive mouse T cell hybridomas. Finally, the monoclonal antibody, L243 binds to the alpha chain of HLA-DR, and was able to block T cell activation by SEA without blocking SEA binding. These data support the model whereby HLA-DR has two binding sites for SEA. A low affinity site, present on the alpha chain, is required for T cell stimulation by the superantigen, but is insufficient to mediate toxin binding. High affinity binding of HLA-DR to SEA occurs solely through residues on the beta chain, including both histidine 81 and aspartate 76.
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PMID:Functional activity of staphylococcal enterotoxin A requires interactions with both the alpha and beta chains of HLA-DR. 912 63

Superantigens are microbial products which bind both to the TCR beta-chain and, with moderate affinity, to MHC class II molecules. Class II-bearing cells bind the superantigen and present the superantigen to T cells expressing certain TCR beta-chain variable region alleles. We have previously reported that the superantigen staphylococcal enterotoxin B (SEB) binds with moderate affinity to the protein p85 expressed on COS-1, an African Green Monkey kidney fibroblast-like cell line. In the present report we carry out a structural analysis to examine the basis for the interaction of superantigen to p85. We show that SEC1, SEC2, and SEC3 also bind to p85 based on inhibition of the binding of radiolabeled SEB. On the other hand, SEA, SED, SEE and toxic shock syndrome toxin-1 do not exhibit detectable binding. In an effort to characterize the structural basis for the SEB binding to p85, we have generated both amino- and carboxy-terminal truncations of SEB expressed as fusion proteins with the maltose-binding protein of Escherichia coli. Our results show that the full-length SEB fusion protein and a truncation missing the 81 amino-terminal amino acids both compete successfully with native SEB for binding. On the other hand, carboxy-terminal truncations in which 19 or 34 residues are deleted both fail to compete for binding. These results are consistent with results which show that monoclonal anti-SEB antibodies specific for carboxy-terminal determinants block SEB binding to p85, but an amino-terminal mAb fails to exhibit any alteration in binding. These results suggest that residues at or near the carboxy-terminus of SEB play a role in binding to p85.
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PMID:Structural basis for the interaction of superantigen with the alternative superantigen-binding receptor p85. 922 68

Because of the low basal output, measurement of acetylcholine (ACh) release from enteric neurons usually requires cholinesterase inhibition, a condition which is known to interfere with feed-back mechanisms regulating ACh release. In this study, we resorted to a highly sensitive HPLC-ED method to determine the minimum requirement of physostigmine to achieve reliable quantitation of spontaneous endogenous ACh overflow from the guinea-pig isolated colon. Furthermore, in order to assess the degree of interference by physostigmine with cholinergic function, we assessed the effect of scopolamine and oxotremorine (in the presence of physostigmine) on spontaneous ACH overflow (to detect the presence of autoreceptors) and also measured the efficiency of the peristaltic reflex with different physostigmine concentrations. Spontaneous endogenous ACh overflow was detectable only with physostigmine concentrations > or = 10 nM. ACh overflow increased with increasing physostigmine concentrations (10 nM-10 microM range). Scopolamine significantly enhanced the facilitatory effect of physostigmine concentrations > or = 10 nM; conversely, oxotremorine inhibited ACh overflow. Peristaltic efficiency was not significantly affected by physostigmine concentrations < or = 300 nM. In conclusion, this modified HPLC-ED method allows ACh detection with minimal physostigmine concentrations (10-30 nM), which do not interfere with peristaltic activity, do not saturate autoreceptor feed-back mechanisms and therefore improve the assessment of cholinergic function in colonic enteric neurons.
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PMID:Acetylcholine detection by a modified HPLC-ED method improves the assessment of cholinergic function in the myenteric plexus of the guinea-pig colon. 929 79

This study compared specific phenotypic and potential virulence characteristics of Staphylococcus aureus isolates from invasive infections and nasal carriers. Three hundred and sixty isolates were studied; 154 from septicaemia (69 line associated, 85 non-line), 79 from continuous ambulatory peritoneal dialysis (CAPD) peritonitis, 64 from bone/joint infections and 64 from healthy nasal carriers. The isolates were tested for production of enterotoxins (SE) A, B, C or E, toxic shock syndrome toxin-1 (TSST-1) protein A, and also for lipolytic, proteolytic, fibrinolytic and haemolytic activities. In addition phage typing, crystal violet reaction, urease and galactose breakdown were studied. Seventy-one percent of isolates were enterotoxigenic. Production of SEA was significantly lower amongst the bone/joint isolates. Production of SEB, was lower among the control group compared with CAPD, bone/joint, and non-line septicaemia isolates. SEE production was higher among the bone/joint isolates compared with the CAPD and non-line septicaemias and production of TSST-1 was significantly higher among nasal isolates compared with isolates causing infection. Almost all of the isolates were lipolytic, with highest activity amongst nasal and bone/joint isolates. Fibrinolytic activity was similar in the five groups of isolates. Proteolytic activity ranged from 35 to 62% of isolates with the lowest frequency among septicaemia isolates. In all, 80-90% of isolates were haemolytic, although CAPD isolates were less likely to be haemolytic. Isolates from the control and CAPD group more frequently belonged to phage group I. TSST-1 does not appear to be an important requirement for invasive infections, but SEB may be. Proteolysis and intensity of lipolysis appear to be less important in septicaemia, and haemolysis may not be important in CAPD peritonitis.
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PMID:Comparative phenotypic characteristics of Staphylococcus aureus isolates from line and non-line associated septicaemia, CAPD peritonitis, bone/joint infections and healthy nasal carriers. 951 32


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