Isolation and Characterization of Messenger RNAs for Seed Lectin and Kunitz Trypsin Inhibitor in Soybeans

Vodkin, Lila O.

doi:10.1104/pp.68.3.766

Cited by 45 publications

(20 citation statements)

References 29 publications

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“…Polysomes from developing maize kernels of inbred line SSA419 were isolated 26 d after pollination according to the procedure of Vodkin (22). Polysome bound mRNAs were in vitro translated in a Wheat Germ System (BRL) according to the procedure of the supplier and using [3H]proline as label.…”

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confidence: 99%

Primary Structure of a Proline-Rich Zein and Its cDNA

Wang¹,

Esen²

1986

Plant Physiol.

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Eighty-five cDNA clones for 'y-zein (proline-rich zein) from a cDNA expression library were isolated using specific antibody and cDNA probes. Nucleotide sequences of seven independent clones were determined and found to be identical in regions where they overlapped. The primary structure of the mature protein, determined from the sequence of one near full-length clone, consists of 204 amino acids. It has a molecular weight of 21,824 daltons, about 5 kilodaltons less than that estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

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confidence: 99%

Primary Structure of a Proline-Rich Zein and Its cDNA

Wang¹,

Esen²

1986

Plant Physiol.

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“…The lectin and trypsin inhibitor antibodies were also affinity purified and have been used many times in conventional immunoprecipitation experiments and western blots (Vodkin, 1981;Jofuku and Goldberg, 1989;Lindstrom et al, 1990) showing their specificity.…”

Section: Polyclonal Antibodies Generated To Synthetic Peptides Represmentioning

confidence: 99%

Direct Detection of Transcription Factors in Cotyledons during Seedling Development Using Sensitive Silicon-Substrate Photonic Crystal Protein Arrays

Jones¹,

Tan

Shamimuzzaman

et al. 2015

Plant Physiology

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Transcription factors control important gene networks, altering the expression of a wide variety of genes, including those of agronomic importance, despite often being expressed at low levels. Detecting transcription factor proteins is difficult, because current high-throughput methods may not be sensitive enough. One-dimensional, silicon-substrate photonic crystal (PC) arrays provide an alternative substrate for printing multiplexed protein microarrays that have greater sensitivity through an increased signal-to-noise ratio of the fluorescent signal compared with performing the same assay upon a traditional aminosilanized glass surface. As a model system to test proof of concept of the silicon-substrate PC arrays to directly detect rare proteins in crude plant extracts, we selected representatives of four different transcription factor families (zinc finger GATA, basic helix-loop-helix, BTF3/ NAC [for basic transcription factor of the NAC family], and YABBY) that have increasing transcript levels during the stages of seedling cotyledon development. Antibodies to synthetic peptides representing the transcription factors were printed on both glass slides and silicon-substrate PC slides along with antibodies to abundant cotyledon proteins, seed lectin, and Kunitz trypsin inhibitor. The silicon-substrate PC arrays proved more sensitive than those performed on glass slides, detecting rare proteins that were below background on the glass slides. The zinc finger transcription factor was detected on the PC arrays in crude extracts of all stages of the seedling cotyledons, whereas YABBY seemed to be at the lower limit of their sensitivity. Interestingly, the basic helix-loop-helix and NAC proteins showed developmental profiles consistent with their transcript patterns, indicating proof of concept for detecting these low-abundance proteins in crude extracts.

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“…In vivo and in vitro studies of lectin synthesis in soybean (Glycine max) (14), field bean (Vicia faba) (4), pea (Pisum sativum) (5,6), french bean (Phaseolus vulgaris) (1), castor bean (Ricinus communis) (10), rice (Oryza sativa), and wheat (Triticum aestivum) (9) have shown that these lectins are all synthesized as higher mol wt precursors which are subsequently processed at the co-and/or posttranslational levels.…”

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Biosynthesis of the Snowdrop (Galanthus nivalis) Lectin in Ripening Ovaries

Damme¹,

Peumans²

1988

Plant Physiol.

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The biosynthesis and processing of the Galanthus nivalis agglutinin were studied in vivo in ripening snowdrop ovaries. Using labeling and pulse chase labeling experiments it could be demonstrated that the snowdrop lectin is synthesized as a precursor of relative molecular weight (Mr) 15,000 which is posttranslationally converted into the authentic lectin polypeptide Of Mr 13,000 with a half-life of about 6 hours. Gel filtration of an extract of [3HJleucine labeled ovaries on Sepharose 4B showed that a significant portion of the newly synthesized lectin is associated with the particulate fraction. When the organeliar fraction was fractionated on isopycnic sucrose gradients this lectin banded in the same density region as the endoplasmic reticulum (ER) marker enzyme NADH cytochrome c reductase.Both radioactivity in lectin and in enzyme activity shifted towards a higher density in the presence of 2 miliimolar Mg- Until now most of the information about the biosynthesis and processing of plant lectins has been obtained with seed lectins. In vivo and in vitro studies of lectin synthesis in soybean (Glycine max) (14), field bean (Vicia faba) (4), pea (Pisum sativum) (5, 6), french bean (Phaseolus vulgaris) (1), castor bean (Ricinus communis) (10), rice (Oryza sativa), and wheat (Triticum aestivum) (9) have shown that these lectins are all synthesized as higher mol wt precursors which are subsequently processed at the co-and/or posttranslational levels.Lectins parts of the flower, especially the ovary (E Van Damme, unpublished data). The GNA' is a tetrameric protein built up of four identical subunits of M, 13,000, which does not contain carbohydrate (12).In this paper we report the biosynthesis of the snowdrop lectin in ripening ovaries. We show evidence that this lectin is synthesized in the ER as a precursor of M, 15,000 which is posttranslationally modified. MATERIALS AND METHODSMaterial. Flowering plants of Galanthus nivalis were collected locally. Unless used immediately they were kept in water to prevent wilting. Ovaries were excised immediately before the onset of the experiments.Radioactive Labeling of the Ovaries. Ovaries were cut longitudinally and each half incubated with a small droplet (10 ,l) containing 10 ,Ci [3H]leucine for appropriate times. Five ovaries were used for each experimental condition. In chase experiments ovaries were labeled with [3H]leucine for 2 h, washed free of labeled precursor, and incubated for different times in the presence of 20 Al of a saturated solution of unlabeled leucine. During the experiments ovaries were kept on a parafilm sheet in closed Petri dishes at 20°C. To prevent dessication moistened filter paper was placed below the parafilm sheets on which the ovaries were kept.Tissue Homogenization. After labeling for the appropriate times the ovaries were rinsed with distilled water and blotted dry. Two different media were used for homogenization and isolation of the organelles. Both contained 100 mm Tris-HCl (pH 7.8) and 12% sucrose; medium A contained in addi...

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Isolation and Characterization of Messenger RNAs for Seed Lectin and Kunitz Trypsin Inhibitor in Soybeans

Cited by 45 publications

References 29 publications

Primary Structure of a Proline-Rich Zein and Its cDNA

Primary Structure of a Proline-Rich Zein and Its cDNA

Direct Detection of Transcription Factors in Cotyledons during Seedling Development Using Sensitive Silicon-Substrate Photonic Crystal Protein Arrays

Biosynthesis of the Snowdrop (Galanthus nivalis) Lectin in Ripening Ovaries

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