Production, Characterization, and Applications of Monoclonal Antibodies Reactive with Soybean Nodule Xanthine Dehydrogenase

Triplett, Eric W.; Lending, Craig R.; Gumpf, D. J.; Ware, Carl F.

doi:10.1104/pp.80.4.965

Cited by 7 publications

(4 citation statements)

References 18 publications

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“…Although XDH is not nodule-specific, an elevated level of XDH is present in nodules (14,16). The higher concentration of XDH observed in the uninfected cells implies that these cells may be the site of xanthine hydroxylation for ureide biogenesis in nodules and that xanthine or a precursor to xanthine, rather than uric acid, is the intermediate transported from the infected to the uninfected cells.…”

Section: Discussionmentioning

confidence: 93%

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Localization of xanthine dehydrogenase in cowpea root nodules: implications for the interaction between cellular compartments during ureide biogenesis.

Datta

Triplett

Newcomb

1991

Proc. Natl. Acad. Sci. U.S.A.

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Immunocytochemistry was used to assess the location of xanthine dehydrogenase (EC 1.1.1.204) in the infected region of nodules of cowpea (Vigna unguiculata [L.] Walpers cv. Queen Anne Blackeye). Polyclonal antibodies raised against purified cowpea xanthine dehydrogenase were used to localize this enzyme at the electron microscopic level. Sparse nonspecific labeling was observed after treatment of nodule sections with preimmune serum. Although immune serum cross-reacted with the ground cytoplasm of both infected and uninfected cells, significantly more labeling was observed in the uninfected cells. No labeling above background was observed in peroxisomes, mitochondria, proplastids, endoplasmic reticulum, cytoplasmic or peribacteroid membranes, peribacteroid spaces, or bacteroids. The enzyme is soluble and not present in any organelle or membrane. The greater concentration of xanthine dehydrogenase in the uninfected cells suggests that xanthine or a precursor to xanthine, rather than uric acid, is the intermediate that moves from infected to uninfected cells during ureide biogenesis.

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Section: Discussionmentioning

confidence: 93%

“…The plants were grown as described (13). The polyclonal antibodies reactive with cowpea XDH used in these experiments were prepared and characterized as described (14)(15)(16). The antibodies used in this study react specifically with XDH (15).…”

mentioning

confidence: 99%

Localization of xanthine dehydrogenase in cowpea root nodules: implications for the interaction between cellular compartments during ureide biogenesis.

Datta

Triplett

Newcomb

1991

Proc. Natl. Acad. Sci. U.S.A.

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“…The 33-kD subunit of uricase has been demonstrated to be identical to the nodule-specific nodulin 35 (3,15). On the basis of immunological studies, xanthine dehydrogenase from soybean has been suggested by Nguyen et al (20) to be nodule-specific, while Triplett et al (29) found significant cross-reactivity in roots, stems, and leaves using monoclonal antibodies raised against soybean nodule xanthine dehydrogenase. However, as has been demonstrated with glutamine synthetase from Phaseolus (8,13), the expression of different forms of an enzyme can be nodule-specific.…”

mentioning

confidence: 98%

Appearance of Purine-Catabolizing Enzymes in Fix⁺ and Fix⁻ Root Nodules on Soybean and Effect of Oxygen on the Expression of the Enzymes in Callus Tissue

Larsen¹,

Jochimsen²

1987

Plant Physiol.

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The appearance of enzymes involved in the formation of ureides, allantoin, and allantoic acid, from inosine 5'-monophosphate was analyzed in developing root nodules of soybean (Glycine max). Concomitant with development of effective nodules, a substantial increase in specific activities of the enzymes 5'-nucleotidase (35-fold), purine nucleosidase (10-fold), xanthine dehydrogenase (25-fold), and uricase (200-fold), over root levels was observed. The specific activity of allantoinase remained constant during nodule development. With ineffective nodules the activities were generally lower than in effective nodules; however, the activities of 5'-nucleotidase and allantoinase were 2-fold higher in ineffective nodules unable to synthesize leghemoglobin than in effective nodules. Since the expression of uricase has been shown to be regulated by oxygen (K Larsen, BU Jochimsen 1986 EMBO J 5: 15-19), the expression of the remaining enzymes in the purine catabolic pathway were tested in response to variations in 02 concentration in sterile soybean callus tissue.

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“…Reports of the use of monoclonal antibodies to nodule-specific plant proteins are sparse. Brewin et al (2) have produced MAbs to a peribacteroid membrane glycoprotein and Triplett et al (22) described MAbs to xanthine dehydrogenase, a key enzyme in the synthesis of ureides. Ureides represent the major form of fixed N2transported from the nodule to the shoots of many legumes (10).…”

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confidence: 99%

Production and Characterization of Monoclonal Antibodies against Aspartate Aminotransferase-P₂ from Lupin Root Nodules

Jones¹,

Reynolds²,

Jones³

et al. 1990

Plant Physiol.

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Twenty-one monoclonal antibodies were raised against the aspartate aminotransferase-P2 isoenzyme from root nodules of Lupinus angustifolius [L.] cv Uniharvest. Induction of this isoenzyme is positively correlated with the onset of N2 fixation in effective root nodules and is associated with the assimilation of ammonia by the plant in the Rhizobium-legume symbiosis. The monoclonal antibodies produced were all of the IgG class, recognized five different epitopes on the protein, and represented greater than 90% of the available epitopes. These epitopes were not unique to lupin nodule aspartate aminotransferase-P2 but were shown to be present on the enzyme from tobacco leaves and potato. Four of the epitopes were conformational with a fifth epitope recognized by the appropriate monoclonals in both its native and denatured forms. None of the monoclonal antibodies produced reacted with Rhizobium lupini NZP2257 extracts. Antibodies against two epitopes showed some cross-reaction with the constitutive aspartate aminotransferase-P, isoenzyme also found in lupin root nodules. However, affinity of these monoclonals for AAT-P1 was three orders of magnitude lower than for AAT-P2. Monoclonals against the other epitopes appeared to be specific for aspartate aminotransferase-P2.MAbs' have proven to be useful diagnostic reagents with many applications in the study of tissue of legumes. AAT-P2 is nodule specific and induced during rhizobial infection of roots concomitant with the onset of N2 fixation and is associated with the proplastid fraction (1). Both enzymes have been separated, purified, and characterized enzymically (13). This is the first report of monoclonal antibodies prepared against plant AAT, although there is a recent report of a polyclonal antibody against an AAT isoform from Panicum maximum (1 1). Polyclonal antibodies have been produced against animal enzymes (17) to study the evolution rates of vertebrate aminotransferases, but this was not extended to plants (17). MAbs have been produced against porcine mitochondrial AAT (20). This paper reports the production and characterization ofa range ofMAbs generated against AAT-P2 MATERIALS AND METHODS Plant Growth and Rhizobium InoculationLupin seeds (Lupinus angustifolius [L] cv Uniharvest) were purchased from Dalgety N.Z. Ltd. Seeds were surface-sterilized, germinated on agar, and transferred to sterile pumice troughs before inoculation with Rhizobium lupini NZP2257. Plant/Rhizobium cultures were grown either in glass houses during summer (day length 2 12 h) or in a controlled environment growth cabinet with a day length of 12 h and a day/ night temperature regime of 24°C/2 1°C (14). Purfication of AAT-P2Nodules (18-21 d old) were harvested and homogenized in two volumes of 50 mM Tris (pH 8.0) containing 0.4 M sucrose and 50 ,ug/mL pyridoxal phosphate. AAT-P, and AAT-P2 were separated and partially purified by ammonium sulfate fractionation, gel filtration on Sepharose CL-6B, and ion exchange chromatography on DEAE Sepharose essentially as described prev...

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Production, Characterization, and Applications of Monoclonal Antibodies Reactive with Soybean Nodule Xanthine Dehydrogenase

Abstract: ABSTRACT

Cited by 7 publications

References 18 publications

Localization of xanthine dehydrogenase in cowpea root nodules: implications for the interaction between cellular compartments during ureide biogenesis.

Localization of xanthine dehydrogenase in cowpea root nodules: implications for the interaction between cellular compartments during ureide biogenesis.

Appearance of Purine-Catabolizing Enzymes in Fix⁺ and Fix⁻ Root Nodules on Soybean and Effect of Oxygen on the Expression of the Enzymes in Callus Tissue

Production and Characterization of Monoclonal Antibodies against Aspartate Aminotransferase-P₂ from Lupin Root Nodules

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Production, Characterization, and Applications of Monoclonal Antibodies Reactive with Soybean Nodule Xanthine Dehydrogenase

Abstract: ABSTRACT

Cited by 7 publications

References 18 publications

Localization of xanthine dehydrogenase in cowpea root nodules: implications for the interaction between cellular compartments during ureide biogenesis.

Localization of xanthine dehydrogenase in cowpea root nodules: implications for the interaction between cellular compartments during ureide biogenesis.

Appearance of Purine-Catabolizing Enzymes in Fix+ and Fix− Root Nodules on Soybean and Effect of Oxygen on the Expression of the Enzymes in Callus Tissue

Production and Characterization of Monoclonal Antibodies against Aspartate Aminotransferase-P2 from Lupin Root Nodules

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Product

Resources

About

Appearance of Purine-Catabolizing Enzymes in Fix⁺ and Fix⁻ Root Nodules on Soybean and Effect of Oxygen on the Expression of the Enzymes in Callus Tissue

Production and Characterization of Monoclonal Antibodies against Aspartate Aminotransferase-P₂ from Lupin Root Nodules