Nucleotide sequence of cDNA coding for saporin‐6, a type‐1 ribosome‐inactivating protein from <i>Saponaria officinalis</i>

Benatti, Luca; Saccardo, Maria Beatrice; Dani, Maria; Nitti, Gianpaolo; Sassano, Marica; Lorenzetti, Rolando; Lappi, Douglas A.; Soria, Marco R.

doi:10.1111/j.1432-1033.1989.tb14951.x

Cited by 56 publications

(23 citation statements)

References 41 publications

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“…This predicted the existence of at least four seed isoforms and, indeed, several different genomic clones were successively identified, confirming the existence of a multigene saporin family [11]. In addition, a leaf cDNA clone had been found to encode a saporin precursor, giving rise to a mature polypeptide of 253 amino acids that differed from SAP-S at 13 amino acid residues [12,13]. In contrast with type II RIPs, type I RIPs are active not only against Abbreviations used : RIP, ribosome-inactivating protein ; SAP, saporin isoform ; α2MR, α2-macroglobulin receptor ; PAP, pokeweed antiviral protein ; RP-HPLC, reversed-phase HPLC ; 3D, three-dimensional ; TFA, trifluoroacetic acid ; TCA, trichloroacetic acid ; CHO, Chinese hamster ovary ; RAP, receptor-associated protein ; LpL, lipoprotein lipase ; RNP, ribonucleoprotein.…”

Section: Introductionmentioning

confidence: 73%

“…An NcoI restriction site could be used instead to select recombinants for the other saporin clones, since sequence 3 lacks this site. The saporin-coding leaf cDNA [12] was mutated to introduce a stop codon before the encoded C-terminal propeptide [13]. The resulting construct, pET-11d-SAP-C, was then fully sequenced to confirm that no changes were introduced during the amplification step.…”

Section: Methodsmentioning

confidence: 99%

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Characterization of a saporin isoform with lower ribosome-inhibiting activity

Fabbrini

Rappocciolo

Carpani

et al. 1997

Biochemical Journal

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We have expressed in Escherichia coli five isoforms of saporin, a single-chain ribosome-inactivating protein (RIP). Translation inhibition activities of the purified recombinant polypeptides in itro were compared with those of recombinant dianthin 30, a less potent and closely related RIP, and of ricin A chain. Dianthin 30, and a saporin isoform encoded by a cDNA from leaf tissue (SAP-C), both had about one order of magnitude lower activity in translation inhibition assays than all other isoforms of saporin tested. We recently demonstrated that saporin extracted from seeds of Saponaria officinalis binds to α2-macroglobulin receptor (α2MR ; also termed low density lipoprotein-receptor-related-protein), indicating a general mechanism of interaction of plant RIPs with the α2MR system [Cavallaro, Nykjaer, Nielsen and Soria (1995) Eur. J. Biochem.

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Section: Introductionmentioning

confidence: 73%

Section: Methodsmentioning

confidence: 99%

Characterization of a saporin isoform with lower ribosome-inhibiting activity

Fabbrini

Rappocciolo

Carpani

et al. 1997

Biochemical Journal

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“…The N-terminal half of JIP60 is related to both type I and type II RIPs, such as the Mirabilisjalapa antiviral protein (Habuka et al, 1989), saporin-6 from Saponaria officinalis (Benatti et al, 1989), and ricin from Ricinuscommunis (Lambet al, 1985) (see Reinbothe et al, 1994, for comparison). Whereas type I RlPs are basic proteins that consist of a single polypeptide chain of 25 to 32 kD (for references, see Bass et al, 1992), type II RlPs contain, in addition to the N-terminal RIP domain, a second domain that confers lectinlike properties to these proteins Pihl, 1973, 1982).…”

Section: General Depression Of Translation: Jip60 Lnactivates Proteinmentioning

confidence: 99%

JIPs and RIPs: the regulation of plant gene expression by jasmonates in response to environmental cues and pathogens.

1994

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“…In this way, we could ensure that the hydrophobic signal peptide would anchor the protein into the membrane, allowing its retention within the early secretory pathway. The selected signal peptide was that of the type I ribosome-inactivating protein saporin (55), containing T22N and A24V to prevent cleavage by signal peptidase without affecting ER import. 6 When we followed the fate of this glycosylated protein (sapRTB) by pulse-chase, it was noticeable that, in contrast to native RTB (Fig.…”

Section: Journal Of Biological Chemistry 33281mentioning

confidence: 99%

Ricin B Chain Targeted to the Endoplasmic Reticulum of Tobacco Protoplasts Is Degraded by a CDC48- and Vacuole-independent Mechanism*

Chamberlain

Marshall

Jolliffe

et al. 2008

Journal of Biological Chemistry

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The B chain of ricin was expressed and delivered to the endoplasmic reticulum of tobacco protoplasts where it disappeared with time in a manner consistent with degradation. This turnover did not occur in the vacuoles or upon secretion. Indeed, several lines of evidence indicate that, in contrast to the turnover of endoplasmic reticulum-targeted ricin A chain in the cytosol, the bulk of expressed ricin B chain was degraded in the secretory pathway.Ricin is a heterodimeric plant protein consisting of a catalytic ribosome-inactivating polypeptide (the A chain, or RTA) 5 disulfide-bonded to a galactose-specific lectin (the B chain, or RTB) (1). In this form, it is able to enter mammalian cells to reach the endoplasmic reticulum (ER) where, following toxin reduction, the RTA subunit is exported to the cytosol in a process that probably exploits some or all phases of the quality control pathway known as ER-associated protein degradation (ERAD) (2, 3). Although a significant proportion of RTA is eventually degraded by proteasomes, a fraction appears to uncouple from this pathway to refold and inactivate substrate ribosomes (4). This inactivation results from a specific depurination of 28 S rRNA at a site essential for the binding of elongation factors during protein synthesis (5). In mammalian cells, the fate of endocytosed RTB is not known. During the biosynthesis of ricin in the producing castor oil plant, the protein initially folds within the ER lumen. However, retro-translocation of RTA is avoided by the translation and ER segregation of an A-B precursor (proricin) that is incompetent for such a step (6). Instead, the ER-sequestered precursor is transported to vacuoles by virtue of a targeting signal that lies in a propeptide linking the two polypeptides. This internal sequence is removed by proteolysis only when proricin reaches the safe haven of storage vacuoles (7). In this way, sensitive plant ribosomes remain undamaged in the wake of large scale synthesis of a highly toxic protein.We have shown previously that in plant cells, in contrast to the fate of proricin, ER-sequestered RTA (rather like RTA reduced from ricin in the mammalian ER) was susceptible to proteasomal degradation following its retro-translocation and deglycosylation in the cytosol (8). As in mammalian cells, however, a fraction of dislocated RTA was able to refold to inhibit protein synthesis. This was the first demonstration of an operational retro-translocation pathway in plant cells (9, 10), and it highlighted the danger to the plant cell of expressing damaged transcripts or prematurely processed proricin. In contrast, when RTA was co-expressed with RTB, where both nascent proteins contained an ER signal peptide, a disulfide-bonded holotoxin was generated and subsequently secreted from the cell (8). The presence, on one or other of the subunits, of the previously characterized vacuolar targeting sequence, directed this holotoxin to vacuoles in a route akin to that of the proricin precursor (7, 11). These findings clearly showed that co-expr...

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Nucleotide sequence of cDNA coding for saporin‐6, a type‐1 ribosome‐inactivating protein from Saponaria officinalis

Cited by 56 publications

References 41 publications

Characterization of a saporin isoform with lower ribosome-inhibiting activity

Characterization of a saporin isoform with lower ribosome-inhibiting activity

JIPs and RIPs: the regulation of plant gene expression by jasmonates in response to environmental cues and pathogens.

Ricin B Chain Targeted to the Endoplasmic Reticulum of Tobacco Protoplasts Is Degraded by a CDC48- and Vacuole-independent Mechanism*

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