Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Intracellular pH has recently been shown to increase during parthenogenetic activation of the porcine oocyte. In the following set of experiments, intracellular pH was monitored during activation and pronuclear development was assessed following activation treatments with calcium, in the absence of calcium, and in oocytes loaded with the calcium chelator BAPTA-AM in calcium-free medium. Intracellular pH increase was not different among groups when treating with 7% ethanol or 50 microM calcium ionophore, or during treatment with thimerosal for 12 or 25 min. Activation with thimerosal (200 microM, 12 min) followed by 8 mM dithiothreitol (DTT, 30 min) resulted in a decreased pronuclear development in calcium-free medium with or without BAPTA-AM loaded oocytes as compared to controls. Activation with 50 microM calcium ionophore resulted in pronuclear development that was different between the calcium-free and BAPTA-AM loaded oocytes in calcium-free medium. Similar incidences of pronuclear formation were observed in all ethanol treatment groups. It was concluded that external calcium as well as large changes in intracellular free calcium are not necessary for the increase in intracellular pH, but normal intracellular calcium signaling is critical for normal levels of pronuclear development. Finally, oocytes were measured for intracellular pH changes for 30 min following subzonal sperm injection. Intracellular pH did not increase, although pronuclear formation was observed 6 hr post SUZI. This suggested that major differences were still present between sperm-induced and parthenogenetic activation of the porcine oocyte.
Mol Reprod Dev 2001 Jun
PMID:Porcine oocyte activation: differing roles of calcium and pH. 1138 59

Understanding the response of tumors to ionizing radiation might potentially lead to improvement in tumor control and patient morbidity. Since the antioxidant status is likely to be linked to radioresponse, its modulation needs to be examined. Therefore, Swiss albino male mice (7-8 weeks old) with Ehrlich solid tumors were irradiated with different doses of gamma rays (0-9 Gy) at a dose rate of 0.0153 Gy/s; and enzymes involved in antioxidant functions were determined in the tumors. Radiation effects in terms of oxidative damage, LDH, nitric oxide and DNA fragmentation were also examined. In tumors, the specific activity of SOD was increased with dose but declined 6 Gy onwards. GST, DTD and GSH showed an almost progressive increase. These enhanced activities might have resulted from the increased protein expression. This possibility was supported by the Western Blot analysis for GST protein. These changes might be closely linked to the radiation-induced oxidative stress as reflected by the enhanced levels of peroxidative damage, DNA fragmentation, LDH activity and nitric oxide levels. These findings may have relevance to radiation therapy of cancer as the elevated antioxidant status of irradiated tumors is likely to limit the effectiveness of radiation dose and adversely affect the therapeutic gain.
Mol Cell Biochem 2001 Jul
PMID:Radiation induced oxidative stress: I. Studies in Ehrlich solid tumor in mice. 1168 24

The effect of regucalcin, a regulatory protein of Ca2+ signaling, on guanosine-5'-triphosphatase (GTPase) activity in isolated rat liver plasma membranes was investigated. GTPase activity was significantly increased by the addition of Ca2+ (25-100 microM) in the enzyme reaction mixture. Such an increase was not seen by other metals (Mg, Co, Zn, Cu, Ni and Mn) with 50 microM. The activatory effect of calcium (50 microM) was significantly decreased by calmodulin (2.5 and 5 microg/ml), indicating that it does not depend on calmodulin. The presence of regucalcin (0.1-0.5 microM) in the enzyme reaction mixture caused a significant increase in GTPase activity. This increase was not significantly enhanced by calcium (50 microM). GTPase activity was significantly increased by dithiothreitol (DTT; 5 mM), a protecting reagent of thiol (SH)-groups, while it was decreased by N-ethylmaleimide (NEM; 5 mM), a modifying reagent of SH-groups. The effect of calcium or regucalcin in increasing GTPase activity was not seen in the presence of NEM. Also, the activatory effect of calcium or regucalcin on GTPase was not seen in the presence of vanadate, an inhibitor of protein phosphorylation, which could inhibit GTPase activity. Moreover, the effect of regucalcin was not seen in the presence of digitonin (0.01%), a solubilizing reagent of membranous lipids, while the effect of calcium was not inhibited by digitonin. The present study demonstrates that regucalcin has an activatory effect on GTPase activity independently of Ca2+ in rat liver plasma membranes.
Mol Cell Biochem 2001 Aug
PMID:Activatory effect of regucalcin on GTPase activity in rat liver plasma membranes. 1169 88

Since the radiation dose tolerance of normal tissues/organs away from the site of tumor influences the success of radiation therapy of cancer, and antioxidant status is likely to be one of the factors to determine the tolerance; the radioresponse of antioxidant enzymes has been examined in the liver as a representative distant organ in the tumor-bearing mice. Swiss albino male mice (7-8 weeks old) with Ehrlich solid tumor in the thigh pad were irradiated with different doses of gamma-radiation (0-9 Gy) at a dose rate of 0.0153 Gy/s and the specific activities of enzymes involved in the free radical metabolism were determined in the liver. Except GST, the activities of SOD, DTD and Gly I as well as the GSH content were found to be higher in the liver of tumor-bearing mice compared to the non-tumor bearing mice. The catalase activity progressively decreased with dose in both the groups of mice. However, the activity was relatively higher in the liver of tumor- bearing mice than the control. Thus, the radioresponse of antioxidant enzymes seemed to be significantly different in the liver of tumor-burdened mice compared to controls. The enhanced activities might be due to relatively more damage caused by radiation. The higher levels of NO* and peroxidative damage in the liver of tumor-bearing mice probably suggest this possibility. These findings of the present work might have some serious implications as the increased radiation-damage of the distant normal organs (due to tumor burden) is likely to adversely affect the therapeutic gain.
Mol Cell Biochem 2001 Aug
PMID:Radiation induced oxidative stress: II studies in liver as a distant organ of tumor bearing mice. 1169 3

The aim of this study was to produce the Bet v 1-related major hazelnut allergen Cor a 1.0401 and variants thereof as recombinant allergens, and to compare their immuno-reactivity with the major hazel pollen allergen using sera of patients whose hazelnut allergy recently was confirmed by double-blind placebo-controlled food challenges (DBPCFC) in a multicenter study. Total RNA was isolated from immature hazelnuts and transcribed into cDNA. Full length coding DNA obtained by PCR-strategy was subcloned into pTYB11 vector and expressed in E. coli ER2566 cells. Native non-fusion target proteins were purified by DTT-induced self-cleavage of the intein-tagged N-terminal fusion proteins. IgE reactivity of the recombinant allergens was tested by enzyme allergosorbent test (EAST), EAST-inhibition, immunoblot-inhibition and histamine release assays. Four recombinant allergens were produced showing deduced amino acid sequence identities among each other of 97-99%, and were considered as variants Cor a 1.0401 (GenBank Accession no.: AF136945), Cor a 1.0402 (AF323973), Cor a 1.0403 (AF323974) and Cor a 1.0404 (AF323975). Cor a 1.0402 and 03 only differed in a C4S exchange. Cor a 1.0404 had a unique proline residue in position 99. Surprisingly, only 63% identity was revealed with hazel pollen Cor a 1. EAST with 43 sera of patients with positive DBPCFC to hazelnut indicated IgE reactivity to Cor a 1.0401 in 95% of the sera, to Cor a 1.0402 in 93%, to Cor a 1.0403 in 91%, and in only 74% of the sera to the proline variant Cor a 1.0404. The allergenic activity of the four variants was confirmed by histamine release assays in 15 hazelnut-allergic patients stimulated with the four variants and controls. Eleven sera were positive with extract from native hazelnut, 13 with rCor a 1.0401, 12 with rCor a 1.0402, 11 with rCor a 1.0403, and only two with rCor a 1.0404 containing the proline exchange. The high IgE binding variant Cor a 1.0401 showed only partial IgE cross-reactivity with pollen Cor a 1. IgE-binding and histamine release capacity led to a concordant ranking of the allergenic activity of the recombinant variants: Cor a 1.0401>Cor a 1.0402 and 03>Cor a 1.0404 (the proline variant). Similar results for Cor a 1.0402 and 03 suggest a minor influence in IgE binding of cysteine in position 4, whereas proline in position 99 appears to be responsible for the decrease in IgE reactivity in Cor a 1.0404. It appears that the epitopes of hazelnut Cor a 1.04 are less related to pollen Cor a 1 than to Bet v 1 from birch pollen. Low IgE binding variants or mutants of Cor a 1.04 are candidate compounds for developing a novel and safe approach of specific immunotherapy of hazelnut allergy.
Mol Immunol 2002 Jan
PMID:Comparison of four variants of a major allergen in hazelnut (Corylus avellana) Cor a 1.04 with the major hazel pollen allergen Cor a 1.01. 1175 Jun 53

Here we report the cloning and characterization of a gene, cypA, from Aspergillus niger that encodes a peptidyl prolyl cis-trans isomerase (PPIase) belonging to the cyclophilin family. Sequencing of both genomic and cDNA clones revealed two ORFs in cypA, one encoding a 19-kDa protein of 174 amino acid residues and the other a 24-kDa protein of 219 amino acid residues, with overall identities of 27-77% to the homologous cyclophilins from prokaryotic and eukaryotic organisms. Expression of the 19-kDa CYPA-(His)(6) in E. coli shows that the purified protein has PPIase activity which is inhibited by cyclosporin A. Northern analysis shows two specific cypA transcripts, the smaller transcript encodes the cytosolic 19-kDa CYPA protein, the larger transcript encodes the putative mitochondrial 24 kDa CYPA protein. The transcript for the cytosolic CYPA is expressed at a higher basal level than that for the mitochondrial protein. The presence of tunicamycin, DTT or cyclosporin A in the medium does not affect the expression level of cypA. Its expression is however slightly induced by heat shock. Growing A. niger mycelium in the presence of cyclosporin A leads to an increase in hyphal branching prior to growth arrest. Overexpression of cypA under the control of its own promoter in A. niger results in increased sensitivity to cyclosporin A, suggesting that cypA encodes the cellular target for cyclosporin A in A. niger.
Mol Genet Genomics 2001 Dec
PMID:The Aspergillus niger cypA gene encodes a cyclophilin that mediates sensitivity to the immunosuppressant cyclosporin A. 1181 Feb 23

Here we report the isolation and characterization of the cypB gene from Aspergillus niger. The cypB gene encodes a protein with a predicted molecular weight of 20.7 kDa, which shows a high degree of identity to the cyclophilin family of peptidyl prolyl cis-trans isomerases (PPIases) from other eukaryotes. The 5' untranslated region of cypB includes three sequences resembling UPREs (unfolded protein response elements). The expression of cypB is upregulated by tunicamycin and DTT, suggesting that at least one UPRE is functional. The CYPB protein also has a 23-amino acid sequence which serves to target the protein to the endoplasmic reticulum (ER), and the ER retention sequence HEEL. CYPB-(His)(6) was expressed in Escherichia coli; the purified protein is capable of isomerizing a substrate peptide in vitro. This is also the first report to show that C-terminal addition of the sequence HEEL is sufficient to ensure retention of the green fluorescent protein (GFP) within the ER.
Mol Genet Genomics 2001 Dec
PMID:The foldase CYPB is a component of the secretory pathway of Aspergillus niger and contains the endoplasmic reticulum retention signal HEEL. 1181 Feb 24

The function of a gene closely linked to nitrate assimilation loci from Chlamydomonas reinhardtii has been investigated. Gene expression analysis shows that its mRNA accumulation is modulated by light, carbon source and adaptation to light/dark cyclic conditions of growth. A full-length cDNA was isolated for the light-regulated transcript, and sequence characterization indicates that it encodes the NADP-malate dehydrogenase from C. reinhardtii (NADP-MDH;Cr). The primary structure of NADP-MDH;Cr is closely related to plant, mossfern and algal NADP-malate dehydrogenases, and shares structural determinants for chloroplast targeting, cofactor binding and catalysis. Sequence conservation extends to the carboxy end of the protein, where plant and mossfern enzymes have two cysteines and an acidic C-terminus with a critical role for regulation of NADP-MDH activity by the thioredoxin/ferredoxin system. Accordingly, incubation with DTT activates NADP-MDH enzyme in cell-free extracts from C. reinhardtii. Like NADP-malate dehydrogenases from two other green algae, the N-terminal extension of NADP-MDH;Cr lacks two thiol residues whose reduction constitutes the rate-limiting step in the activation reaction of plant enzymes. Homology-based 3D modelling of NADP-MDH;Cr, the first structure predicted for NADP-malate dehydrogenase from a lower eukaryote, evidences close positioning of two new cysteines in an accessible region of the protein surface. These results suggest that the algal enzyme has a different arrangement of regulatory disulfide bridges, which might involve the existence of new mechanisms that control functioning of the malate valve, the main system to export reducing power from the chloroplast of plant cells.
Plant Mol Biol 2002 Feb 01
PMID:NADP-malate dehydrogenase from Chlamydomonas: prediction of new structural determinants for redox regulation by homology modelling. 1185 23

The eukaryotic replication protein A (RPA) is a heterotrimeric protein complex. It consists of 70, 32, and 14 kDa subunits that are involved in DNA replication, repair, and genetic recombination. RPA is a 4-cysteine type zinc-finger protein. RPA's zinc-finger domain is not essential for DNA binding activity, but it is involved in the regulation of RPA's DNA binding activity through reduction-oxidation (redox). In this study, we show that yeast RPA's ssDNA binding activity is regulated by redox potential through its subcomplexes of 32 and 14 kDa subunits. In contrast, the subunits' complex, RPA70, formed a stable complex with ssDNA, even under non-reducing conditions. The addition of DTT and H202 had no effect on its DNA binding activity. In RPA70, since the addition of the subcomplexes of the 32 and 14 kDa subunits, it restored the modulating ssDNA binding activity to native RPA's DNA binding activity. These results suggest that the subcomplexes of the 32 and 14 kDa subunits may be involved in the modulating RPA's DNA binding activity through redox change. These studies, therefore, show the novel structure and function relationship of a multiprotein complex in that the role of a specific domain (or one subunit) is regulated by the other subunits.
Mol Cells 2002 Jun 30
PMID:Involvement of subcomplexes of 32 and 14 kDa subunits in RPA's DNA binding activity through redox change. 1213 92

Functional reagents known to bring about the formation of a distinct membrane molecular complex of the subunits of cytochrome b(558) (gp 91(phox) and p22(phox)) were investigated for their influence on the O2- generating capability of liposome incorporated cytochrome b(558) preparations. One, ethyleneglycolbis[sulfo-succinimidylsuccinate], (sulfo-EGS) was found to inhibit O2- generation at concentrations which are known to result in cross-linking the two subunits of cytochrome b(558). Sulfosuccinimidyl [4-azidophenyldithio] propionate, (sulfo-SADP) on the other hand, was found to be a powerful inhibitor of the cytochrome b(558) dependent O2- production at concentrations not able to result in cross linking of the two subunits. Sulfo-SADP inhibits the cytochrome b(558) O2- production 50% at 25 microM, while sulfo-EGS requires 400 microM. For these reagents, the succinimidyl group of sulfo-SADP and sulfo-EGS is the reactive group, which inhibit irreversibly, cytochrome b(558) generation of O2-. Both sulfo-SADP and sulfo-EGS have similar linker arms of 13.9 and 16.1 A, respectively. The difference, accounting for the strong inhibitory profile for sulfo-SADP as compared with sulfo-EGS, resides in the aryl group associated with the sulfo-SADP. The aryl group of sulfo-SADP has been found to be important in directing the specificity of the probe in its inhibition of O2- generation. When the disulfide bond linking the aromatic portion of the probe to the succinimidyl ring is cleaved by DTT (dithiothreitol), the product loses its specificity and has an inhibitory activity with respect to O2- generation comparable to that of sulfo-EGS. The partial protection against the inhibitory influence of sulfo-SADP by NADP(+) indicates that the reagent may interact at the pyridine nucleotide-binding domain of cytochrome b(558). Its low inhibitory titer and its water solubility suggest that sulfo-SADP reacts with a specific amine (the primary reactant for the succinimidyl group) on cytochrome b(558).
J Biochem Mol Biol Biophys 2002 Jun
PMID:Sulfo-SADP (sulfosuccinimidyl[4-azidophenyldithio]propionate) an active site directed reagent inhibiting the NADPH dependent O2- generation of leukocyte cytochrome b(558). 1218 52


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