Linamarase and other ?-glucosidases are present in the cell walls of Trifolium repens L. leaves

Kakes, P.

doi:10.1007/bf00397342

Cited by 69 publications

(43 citation statements)

References 16 publications

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“…This strongly suggests compartmentalization of the cyanoglucosides and the hydrolytic enzymes. Linamarase of white clover (Kakes, 1985) has been shown to be apoplastic and may be located in the epidermis. The tissue and subcellular localization of the cyanoglucosides in white clover are not known.…”

Section: Biochemistrymentioning

confidence: 99%

The cyanogenic polymorphism in Trifolium repens L. (white clover)

1991

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The cyanogenic polymorphism in white clover is controlled by alleles of two independently segregating loci. Biochemical studies have shown that non-functional alleles of the Ac locus, which controls the level of cyanoglucoside produced in leaf tissue, result in the loss of several steps in the biosynthetic pathway. Alleles of the Li locus control the synthesis of the hydrolytic enzyme, linamarase, which is responsible for HCN release following tissue damage. Studies on the selective forces and the distribution of the cyanogenic morphs of white clover are discussed in relation to the quantitative variation in cyanogenesis revealed by biochemical studies. Molecular studies reveal considerable restriction fragment length polymorphism for linamarase homologous genes.

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Section: Biochemistrymentioning

confidence: 99%

The cyanogenic polymorphism in Trifolium repens L. (white clover)

1991

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“…By 6) were subjected to SDS-PAGE. Proteins were either stained by Brilliant Blue G-Colloidal (lanes 1-3) or electroblotted onto nitrocellulose and immunodecorated with affinity purified anti-AH antiserum (lanes [4][5][6]. B, Aliquots (7.5 contrast, PH was absent from the more internal procambial cells, including those housing AH, and from the bundle sheath and cotyledonary storage parenchyma.…”

Section: Immunolocalization Of Ah and Ph By Light Microscopymentioning

confidence: 99%

Tissue and Subcellular Localization of Enzymes Catabolizing (R)-Amygdalin in Mature Prunus serotina Seeds

Swain¹,

Li²,

Poulton³

1992

Plant Physiol.

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In black cherry (Prunus serotina Ehrh.) homogenates, (R)-amygdalin is catabolized to HCN, benzaldehyde, and D-glucose by the sequential action of amygdalin hydrolase, prunasin hydrolase, and mandelonitrile lyase. The tissue and subcellular localizations of these enzymes were determined within intact black cherry seeds by direct enzyme analysis, immunoblotting, and colloidal gold immunocytochemical techniques. Taken together, these procedures showed that the two d-glucosidases are restricted to protein bodies of the procambium, which ramifies throughout the cotyledons. Although amygdalin hydrolase occurred within the majority of procambial cells, prunasin hydrolase was confined to the peripheral layers of this meristematic tissue. Highest levels of mandelonitrile lyase were observed in the protein bodies of the cotyledonary parenchyma cells, with lesser amounts in the procambial cell protein bodies. The residual endosperm tissue had insignificant levels of amygdalin hydrolase, prunasin hydrolase, and mandelonitrile lyase.

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“…4) (13). In common with linamarases from Phaseolus lunatus and Trifolium repens (6,11), cassava linamarase resides in the cell wall (20). The subcellular localizations oflinamarin and the a-hydroxynitrile lyase in cassava are unknown.…”

Section: But Left Peptidementioning

confidence: 99%

Immunocytochemical Localization of Mandelonitrile Lyase in Mature Black Cherry (Prunus serotina Ehrh.) Seeds

Poulton²

1991

Plant Physiol.

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Mandelonitrile lyase (MDL, EC 4.1.2.10), which catalyzes the reversible dissociation of (R)-(+)-mandelonitrile to benzaldehyde and hydrogen cyanide, was purified to apparent homogeneity from mature black cherry (Prunus serotina Ehrh.) seeds by conventional protein purification techniques. This flavoprotein is monomeric with a subunit molecular mass of 57 kilodaltons. Glycoprotein character was shown by its binding to the affinity matrix concanavalin A-Sepharose 4B with subsequent elution by a-methyl-D-glucoside. Upon chemical deglycosylation by trifluoromethanesulfonic acid, the molecular mass was reduced to 50.9 kilodaltons. Two-dimensional gel analysis of deglycosylated MDL revealed the presence of several subunit isoforms of similar molecular mass but differing slightly in isoelectric point. Polyclonal antibodies were raised in New Zealand white rabbits against deglycosylated and untreated MDL. Antibody titers were determined by enzyme linked immunosorbent and dot immunobinding assays, while their specificities were assessed by Western immunoblot analysis. Antibodies raised against untreated Iyase recognized several proteins in addition to MDL. In contrast, antisera raised against deglycosylated MDL were monospecific and were utilized for developmental and immunocytochemical localization studies. SDS-PAGE and immunoblotting analysis of seed proteins during fruit maturation showed that MDL first appeared in seeds shortly after cotyledons began development. In cotyledon cells of mature seeds, MDL was localized primarily in the cell wall with lesser amounts in the protein bodies, whereas in endosperm cells, this labeling pattern was reversed. N-terminal sequence data was gathered for future molecular approaches to the question of MDL microheterogeneity.Cyanogenesis, the production of the respiratory poison HCN3 by biological organisms, was first described in plants almost two Enzyme Assay MDL activity was measured spectrophotometrically by monitoring the conversion of (R,S)-mandelonitrile to benzaldehyde and HCN by the increase in absorbance at 249.6 nm (3 1). Protein EstimationProtein content was estimated by the Bradford procedure (2) with BSA serving as standard. Deglycosylation of MDLChemical deglycosylation of MDL was performed using TFMS as described by Edge et al. (5). The deglycosylated protein was recovered by extraction into aqueous phase followed by overnight dialysis against 4 L of 2 mm pyridine acetate buffer, pH 5.5. The dialyzed preparation was concentrated by ultrafiltration (Amicon Minicon) to approximately 1 mL and stored at -20°C. Generation of Polyclonal Antibodies against MDLBefore immunization ofanimals, homogeneous MDL, with or without prior deglycosylation, was subjected to two-dimensional PAGE (IEF range pH 4-6.5, followed by 10% SDS-PAGE). Areas containing MDL were excised and electroeluted into 10 mm Tris-acetate buffer (pH 6.8 LOCALIZATION OF PRUNUS SEROTINA MANDELONITRILE LYASE alkaline phosphatase-conjugated goat anti-rabbit IgG (diluted 1:1000 with blocking solution) for ...

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Linamarase and other ?-glucosidases are present in the cell walls of Trifolium repens L. leaves

Cited by 69 publications

References 16 publications

The cyanogenic polymorphism in Trifolium repens L. (white clover)

The cyanogenic polymorphism in Trifolium repens L. (white clover)

Tissue and Subcellular Localization of Enzymes Catabolizing (R)-Amygdalin in Mature Prunus serotina Seeds

Immunocytochemical Localization of Mandelonitrile Lyase in Mature Black Cherry (Prunus serotina Ehrh.) Seeds

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