Purification and characterization of a glutathione S-transferase Omega in pig: evidence for two distinct organ-specific transcripts

2001 ◽  
Vol 358 (1) ◽  
pp. 257-262 ◽  
Author(s):  
Patrick ROUIMI ◽  
Patricia ANGLADE ◽  
Anne BENZEKRI ◽  
Philippe COSTET ◽  
Laurent DEBRAUWER ◽  
...  
Keyword(s):  
Peptide Sequencing ◽  
Cross Reactivity ◽  
Glutathione S Transferase ◽  
Gene Encoding ◽  
Simple Method ◽  
Organ Specific ◽  
Cytosolic Glutathione ◽  
A Subunit ◽  
Internal Peptide

A cytosolic glutathione S-transferase (GST, EC 2.5.1.18) from the recently characterized Omega class [GSTO; Board et al. 2000, J. Biol. Chem. 275, 24798–24806] has been identified in pig organs. It was found widely distributed in the different tissues investigated and especially abundant in liver and muscle. The hepatic enzyme has been purified to homogeneity by using its selective affinity for S-hexylglutathione over GSH, thus providing a simple method to isolate mammalian GSTO. The dimeric protein has a subunit molecular mass of 27328Da as measured by electrospray ionization MS. Internal peptide sequencing and complete cDNA sequencing revealed strong similarities with its human recombinant orthologue and two rodent GST-like proteins with the ability to catalyse the GSH-dependent reduction of dehydroascorbate. Additional similarities, including the presence of a specific N-terminal extension and of immunological cross-reactivity, support the results. Moreover, this gene encoding GSTO generates two organ-specific transcripts, suggesting transcriptional mechanisms with a significance that is as yet uncharacterized.

scholarly journals Molecular Characterization of the sucB Gene Encoding the Immunogenic Dihydrolipoamide Succinyltransferase Protein of Bartonella vinsonii subsp. berkhoffii and Bartonella quintana

2003 ◽  
Vol 71 (8) ◽  
pp. 4818-4822 ◽  
Author(s):  
Robert D. Gilmore, ◽  
Amber M. Carpio ◽  
Michael Y. Kosoy ◽  
Kenneth L. Gage
Keyword(s):  
Q Fever ◽  
Clinical Manifestations ◽  
Cross Reactivity ◽  
Emerging Pathogens ◽  
Expression Library ◽  
Gene Encoding ◽  
Bartonella Quintana ◽  
Genomic Expression ◽  
Dihydrolipoamide Succinyltransferase

ABSTRACT Members of the genus Bartonella have historically been connected with human disease, such as cat scratch disease, trench fever, and Carrion's disease, and recently have been recognized as emerging pathogens causing other clinical manifestations in humans. However, because little is known about the antigens that elicit antibody production in response to Bartonella infections, this project was undertaken to identify and molecularly characterize these immunogens. Immunologic screening of a Bartonella vinsonii subsp. berkhoffii genomic expression library with anti-Bartonella antibodies led to the identification of the sucB gene, which encodes the enzyme dihydrolipoamide succinyltransferase. Antiserum from a mouse experimentally infected with live Bartonella was reactive against recombinant SucB, indicating the mounting of an anti-SucB response following infection. Antigenic cross-reactivity was observed with antiserum against other Bartonella spp. Antibodies against Coxiella burnetti, Francisella tularensis, and Rickettsia typhi also reacted with our recombinant Bartonella SucB. Potential SucB antigenic cross-reactivity presents a challenge to the development of serodiagnostic tests for other intracellular pathogens that cause diseases such as Q fever, rickettsioses, brucelloses, tularemia, and other bartonelloses.

scholarly journals Immunological Characterization of a 36kD Polypeptide in Peanuts (Arachis hypogaea L.)

1991 ◽  
Vol 18 (1) ◽  
pp. 11-15 ◽  
Author(s):  
E. H. Shokarii ◽  
A. Esen ◽  
R. W. Mozingo
Keyword(s):  
Time Course ◽  
High Titer ◽  
Cross Reactivity ◽  
Protein Deposition ◽  
Arachis Hypogaea L ◽  
A Subunit ◽  
Whole Seed ◽  
Immunological Assays ◽  
Peanut Proteins

Abstract Antiserum against a 36 kD polypeptide (a subunit of arachin) that occurs in poor blanching peanuts was prepared. This antiserum was of high titer but was not monospecific for the homologous 36 kD anitgen; it showed cross-reactivity with other peanut proteins. The antiserum was adsorbed by the whole seed protein fraction of a good blanching cultivar to render it monospecific. The adsorbed antiserum could accurately differentiate the protein extracts from good blanching and poor blanching peanuts when tested by the enzyme linked immounosorbent assay (ELISA) and by immunoblotting. Time course studies of protein deposition during seed development revealed that the 36 kD polypeptide was present from the early stages of embryogenesis. In addition, it was present in all parts of the cotyledon and not limited to the surface of seed. Although the function of the 36 kD polypeptide is not known, our results indicate that its presence can be detected uising immunological assays.

PAPS-Reductase from Escherichia coli: Characterization of the Enzyme as Probe for Thioredoxins

1987 ◽  
Vol 42 (1-2) ◽  
pp. 93-102 ◽  
Author(s):  
Jens D . Schwenn ◽  
Ulrich Schriek
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Malate Dehydrogenase ◽  
Cross Reactivity ◽  
Enzymatic Assay ◽  
Fructose Bisphosphatase ◽  
Simple Method ◽  
Bacterial Enzyme ◽  
Eukaryotic Organisms

Abstract PAPS-reductase from Escherichia coli was em ployed to detect thioredoxins from pro- and eukaryotic organisms. A simple method for the isolation of this enzyme and properties of the enzymatic assay were described. A comparison betw een thioredoxins detected by the PAPS-reductase and the Fructose-bisphosphatase or NADP malate dehydrogenase was used to assess the validity of the test. The high cross-reactivity of the bacterial enzyme was useful in the purification of heterologous thioredoxins from spinach, Synechococcus, and Saccharomyces cerevisiae.

scholarly journals Identification and Characterization of Shiga Toxin Type 2 Variants in Escherichia coli Isolates from Animals, Food, and Humans

2008 ◽  
Vol 74 (18) ◽  
pp. 5645-5652 ◽  
Author(s):  
Jie Zheng ◽  
Shenghui Cui ◽  
Louise D. Teel ◽  
Shaohua Zhao ◽  
Ruby Singh ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Gene Encoding ◽  
Intestinal Mucus ◽  
One Step ◽  
A Subunit ◽  
Pcr Method ◽  
Identification And Characterization

ABSTRACT There is considerable heterogeneity among the Shiga toxin type 2 (Stx2) toxins elaborated by Shiga toxin-producing Escherichia coli (STEC). One such Stx2 variant, the Stx2d mucus-activatable toxin (Stx2dact), is rendered more toxic by the action of elastase present in intestinal mucus, which cleaves the last two amino acids of the A2 portion of the toxin A subunit. We screened 153 STEC isolates from food, animals, and humans for the gene encoding Stx2dact by using a novel one-step PCR procedure. This method targeted the region of stx 2dact that encodes the elastase recognition site. The presence of stx 2dact was confirmed by DNA sequencing of the complete toxin genes. Seven STEC isolates from cows (four isolates), meat (two isolates), and a human (one isolate) that carried the putative stx 2dact gene were identified; all were eae negative, and none was the O157:H7 serotype. Three of the isolates (CVM9322, CVM9557, and CVM9584) also carried stx 1, two (P1332 and P1334) carried stx 1 and stx 2c, and one (CL-15) carried stx 2c. One isolate, P1130, harbored only stx 2dact. The Vero cell cytotoxicities of supernatants from P1130 and stx 1 deletion mutants of CVM9322, CVM9557, and CVM9584 were increased 13- to 30-fold after treatment with porcine elastase. Thus, Stx2dact-producing strains, as detected by our one-step PCR method, can be isolated not only from humans, as previously documented, but also from food and animals. The latter finding has important public health implications based on a recent report from Europe of a link between disease severity and infection with STEC isolates that produce Stx2dact.

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