Oligosaccharide Side Chains of Glycoproteins that Remain in the High-Mannose Form Are Not Accessible to Glycosidases

Faye, Loı̈c; Johnson, Kenneth D.; Chrispeels, Maarten J.

doi:10.1104/pp.81.1.206

Cited by 29 publications

(10 citation statements)

References 24 publications

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“…Obtained extracts were subjected to the separation on an affinity column with immobilized lectin. A common feature of plant seed glycoproteins is the presence of oligosaccharide chains having an affinity for the plant lectin ConA [35,36]. Therefore, this was used to enrich the fraction of barley and Arabidopsis high-mannose and hybrid-types of N-glycoproteins having affinity for this kind of lectin.…”

Section: Resultsmentioning

confidence: 99%

The Combination of Lectin Affinity Chromatography, Gel Electrophoresis and Mass Spectrometry in the Study of Plant Glycoproteome: Preliminary Insights

2011

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The study of plant proteome is important because proteins and glycoproteins are involved in many crucial biological processes that could enhance the agronomical and nutritional value of crops and food products. The analysis of model organisms (e.g., Arabidopsis thaliana) is a popular means to aid in understanding many plant features. Proteomics has the potential to contribute to cereal science, mainly through the description of grain quality by elucidating the ways in which the genes are expressed during grain filling under particular environmental conditions. Since barley grain composition (i.e., its protein and glycoprotein analysis) is important for the brewing industry, human and animal nutrition, plant breeding or cultivar identification, the profiles of glycoproteins released from barley were one of the aims of this paper. This work combines affinity chromatography with 1D gel electrophoresis followed by mass spectrometry for the characterization of modified plant proteins. Lectin chromatography with Concanavalin A (ConA) was integrated to the procedure for the selective isolation of N-glycoproteins. For the verification of the universality of a chosen approach, proteins were extracted by deionized water from two different model plant materials-barley (Hordeum vulgare) and A. thaliana. Our attention was focused on the glycoproteins present in both barley grains and grains after the malting procedure and Arabidopsis seedlings. The proteins retained in an affinity column were separated by SDS-PAGE, excised, digested by trypsin and studied by MALDI-TOF MS. SDS gels revealed the differences between ConA non-bound and bound fractions. The majority of proteins were found in non-bound fractions. Several important glycoproteins were confirmed in studied materials: barley grain (serine carboxypeptidase; alphaamylase inhibitor BMAI-1), barley malt (e.g., b-D-xylosidase) and Arabidopsis (b-D-xylosidase, peroxidase, etc.).

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Section: Resultsmentioning

confidence: 99%

The Combination of Lectin Affinity Chromatography, Gel Electrophoresis and Mass Spectrometry in the Study of Plant Glycoproteome: Preliminary Insights

2011

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“…The binding of ConA to plant glycoproteins is mediated by high-mannose oligosaccharide sidechains (Faye et al 1986). Secreted proteins were chromatographed on small ConA-Sepharose columns, and elution was performed with a stepwise increase in hapten sugar concentration.…”

Section: Secreted Proteins Have High-mannose Glycans and Complex-typementioning

confidence: 99%

The role of high-mannose and complex asparagine-linked glycans in the secretion and stability of glycoproteins

Driouich

Gonnet

Makkie

et al. 1989

Planta

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Suspension-cultured cells of sycamore (Acer pseudoplatanus L.) secrete a number of acid hydrolases and other proteins that have both highmannose and complex asparagine-linked glycans. We used affinity chromatography with concanavalin A and an antiserum specific for complex glycans in conjunction with in vivo-labeling studies to show that all of the secreted proteins carry glycans. The presence of complex glycans on secretory proteins indicates that they are passing through the Golgi complex on the way to the extracellular compartment. The sodium ionophore, monensin, did not block the transport of proteins to the extracellular medium, even though monensin efficiently inhibited the Golgi-mediated processing of complex glycans. The inhibition of N-glycosylation by tunicamycin reduced by 76% to 84% the accumulation of newly synthesized (i.e. radioactively labeled) protein that was secreted by the sycamore cells, while cytoplasmic protein biosynthesis was not affected by this antibiotic. However, in the presence of glycoprotein-processing inhibitors, such as castanospermine and deoxymannojirimycin, the formation of complex glycans was prevented but glycoprotein secretion was unchanged. These results support the conclusion that N-linked glycan processing is not necessary for sorting, but glycosylation is required for accumulation of secreted proteins in the extracellular compartment.

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“…A common feature of plant seed glycoproteins is the presence of high-mannose oligosaccharide side chains which have an affinity for the plant lectin concanavalin A (Con A) (Chrispeels, 1984;Faye et al, 1986). To determine if GLB 1 and GLB2 contain mannose residues, embryo proteins were subjected to Con A affinity blotting as described by Faye and Chrispeels (1985).…”

Section: Analysis Of Embryo Proteins By Con a Affinity Blottingmentioning

confidence: 99%

Characterization of embryo globulins encoded by the maizeGlb genes

Kriz¹

1989

Biochem Genet

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Two of the most abundant proteins in maize embryos are saline-soluble, water-insoluble globulins. One is a Mr 63,000 protein encoded by the Glb1 gene and the other is a Mr 45,000 component encoded by the Glb2 gene. Both proteins accumulate to high levels during embryo development and are rapidly degraded during the early stages of seed germination. Amino acid composition analysis indicates that these proteins may serve as storage reserves to provide sources of nitrogen and carbon to the germinating embryo. Amino-terminal sequence analysis of the final Glb1 gene product, GLB1, and its immediate precursor, GLB1', indicates that the latter is proteolytically cleaved near the amino terminus to form GLB1. In addition to these biochemical studies, we describe the identification of a novel maize variant which lacks the protein product of the Glb2 gene.

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Oligosaccharide Side Chains of Glycoproteins that Remain in the High-Mannose Form Are Not Accessible to Glycosidases

Cited by 29 publications

References 24 publications

The Combination of Lectin Affinity Chromatography, Gel Electrophoresis and Mass Spectrometry in the Study of Plant Glycoproteome: Preliminary Insights

The Combination of Lectin Affinity Chromatography, Gel Electrophoresis and Mass Spectrometry in the Study of Plant Glycoproteome: Preliminary Insights

The role of high-mannose and complex asparagine-linked glycans in the secretion and stability of glycoproteins

Characterization of embryo globulins encoded by the maizeGlb genes

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