1993 Fiscal Year Final Research Report Summary
Vesicle transport and processing for precursors of vacuolar proteins.
| Project/Area Number |
04640632
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
植物生理学
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| Research Institution | National Institute for Basic Biology |
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Principal Investigator |
HARA-NISHIMURA Ikuko National Institute for Basic Biology, Department of Cell Biology, Research Associate, 基礎生物学研究所, 助手 (00241232)
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| Co-Investigator(Kenkyū-buntansha) |
HARA-NISHIMOTO Ikuko National Institute for Basic Biology, Department of Cell Biology, Research Assoc (00241232)
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| Project Period (FY) |
1992 – 1993
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| Keywords | 2S Albumin / Castor bean / Protein body / Proprotein precursors / Seed proteins / Transport vesicles / Vacuolar processing / Vacuoles |
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| Research Abstract |
Cell fractionation of pulse-chase-labeled developing pumpkin cotyledons demonstrated that proprotein precursors are transported from the endoplasmic reticulum to dense vesicles and then to the vacuoles. Proproteins of various vacuolar proteins including 2S albumin are post-translationally processed into mature forms by the action of a unique vacuolar processing enzyme. The primary structure of the precursor to pumpkin 2S albumin has been deduced from the nucleotide sequence of an isolated cDNA insert. Post-translational cleavages occur on the C-terminal sides of sparagine residues 35 and 74, which are conserved among precursors to 2S albumin from different plants. Hydropathy analysis revealed that the two asparagine residues are located in the hydrophilic regions of pro2S albumin. These findings suggest that a vacuolar processing enzyme can recognize exposed asparagine residues on the molecular surface of pro2S albumin and cleave the peptide bond on the C-terminal side of each asparagi
… More
ne residue to produce mature 2S albumin in the vacuoles.
Immunoelectron micoscopy of the maturing endosperm of castor bean(Ricinus communis)revealed that the vacuolar processing enzyme is selectively localized in the dense vesicles as well as in the vacuolar matrix. This indicates that the vacuolar processing enzyme is transported to vacuoles via dense vesicles as does 2S albumin. The result suggests that the enzyme must be a latent form. To characterize a molecular structure of vacuolar processing enzyme, we isolated a cDNA for the enzyme. Deduced primary structure of 55-kD precursor is 33% identical to a putative cystine proteinase of the human parasite Schistosoma mansoni. The precursor is composed of a signal peptide, a 37-kD active processing enzyme domain, and a propeptide fragment. Although the precursor expressed in Escherichia coli has no vacuolar processing activity, a 36-kD immunopositive protein expressed in E.coli is the active. These results suggest that the activation of the vacuolar processing enzyme requires proteolytic cleavage of the 14-kD C-terminal propeptide fragment of the precursor. Less
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