Posttranslational processing of concanavalin A precursors in jackbean cotyledons

Bowles, D. J., Marcus, S. E., Pappin, D. J., Findlay, J. B., Eliopoulos, E., Maycox, P. R., Burgess, J. (April 1986) Posttranslational processing of concanavalin A precursors in jackbean cotyledons. J Cell Biol, 102 (4). pp. 1284-97. ISSN 0021-9525 (Print)0021-9525 (Linking)

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URL: http://www.ncbi.nlm.nih.gov/pubmed/3958046
DOI: 10.1083/jcb.102.4.1284

Abstract

Metabolic labeling of immature jackbean cotyledons with 14C-amino acids was used to determine the processing steps involved in the assembly of concanavalin A. Pulse-chase experiments and analyses of immunoprecipitated lectin forms indicated a complex series of events involving seven distinct species. The structural relatedness of all of the intermediate species was confirmed by two-dimensional mapping of 125I-tryptic peptides. An initial glycosylated precursor was deglycosylated and cleaved into smaller polypeptides, which subsequently reannealed over a period of 10-27 h. NH2-terminal sequencing of the abundant precursors confirmed that the intact subunit of concanavalin A was formed by the reannealing of two fragments, since the alignment of residues 1-118 and 119-237 was reversed in the final form of the lectin identified in the chase and the precursor first labeled. When the tissue was pulse-chased in the presence of monensin, processing of the glycosylated precursor was inhibited. The weak bases NH4Cl and chloroquine were without effect. Immunocytochemical studies showed that monensin treatment caused the accumulation of immunoreactive material at the cell surface and indicated that the ionophore had induced the secretion of a component normally destined for deposition within the protein bodies. Consideration of the tertiary structure of the glycosylated precursor and mature lectin showed that the entire series of processing events could occur without significant refolding of the initial translational product. Proteolytic events included removal of a peptide from the surface of the precursor molecule that connected the NH2- and COOH-termini of the mature protein. This processing activated the carbohydrate-binding activity of the lectin. The chase data suggest the occurrence of a simultaneous cleavage and formation of a peptide bond, raising the possibility that annealment of the fragments to give rise to the mature subunit involves a transpeptidation event rather than cleavage and subsequent religation.

Item Type: Paper
Uncontrolled Keywords: Amino Acid Sequence Amino Acids/metabolism Carbon Radioisotopes Concanavalin A/biosynthesis/genetics Fabaceae/genetics/metabolism/ultrastructure Kinetics Microscopy, Electron Models, Molecular Molecular Weight Peptide Fragments/analysis Plant Lectins Plants/genetics/ metabolism/ultrastructure Plants, Medicinal Protein Conformation Protein Processing, Post-Translational
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein expression > post-translational modification
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein expression
CSHL Authors:
Communities: CSHL labs > Pappin lab
Depositing User: Matt Covey
Date: April 1986
Date Deposited: 10 Sep 2014 14:13
Last Modified: 10 Sep 2014 14:13
PMCID: PMC2114185
Related URLs:
URI: https://repository.cshl.edu/id/eprint/30771

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