Elderberry (Sambucus Nigra) Bark Contains two Structurally Different Neusac(alpha2,6)Gal/Galnac-Binding Type 2 Ribosome-Inactivating Proteins (original) (raw)
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FEBS Letters, 1998
Analysis of affinity-purified preparations of the fetuin-binding proteins from elderberry bark and fruits revealed besides the previously reported Neu5Ac(K K-2,6)Gal/GalNAcspecific type 2 ribosome-inactivating proteins (RIP) the occurrence of single chain proteins of 22 kDa, which according to their N-terminal amino acid sequence correspond to the second part of the B chain of the respective type 2 RIP. Both proteins are very similar except that the polypeptides of the fruit lectin are 10 amino acid residues longer than these from the bark lectin. Our findings not only demonstrate the occurrence of carbohydratebinding fragments of type 2 RIP but also provide further evidence that type 2 RIP genes give rise to complex mixtures of type 2 RIP/lectins in elderberry.
European Journal of Biochemistry, 1996
The cDNA encoding the NeuAc(a-2,6)Gal/GalNAc binding lectin from elderberry (Samhucus nigrcz) bark (SNAI) was isolated from a cDNA library constructed with mRNA from the bark. Sequence analysis of this lectin cDNA revealed a striking similarity to the previously sequenced type-2 ribosome-inactivating proteins from Ricinus communis and Ahrus precatorius. Molecular modelling of SNAI further indicated that its structure closely resembles that of ricin. Since SNAI strongly inhibits cell-free protein synthesis in a rabbit reticiilocyte lysate it presumably is a type-2 ribosome-inactivating protein. However, SNAI differs from all previously described type-2 ribosome-inactivating proteins by its specificity towards NeuAc(rr-2,6)Gal/GalNAc and its unusual molecular structure.
European Journal of Biochemistry, 1996
The molecular structure of the Sambucus nigra agglutinin V (SNAV), which has been described previously as a type-2 ribosome-inactivating protein called nigrin b, has been studied in detail by analysis of the purified protein combined with cDNA cloning and molecular modelling. Native SNAV is a dimer of two [A-s-s-B] pairs. Hapten inhibition assays indicated that GalNAc is a 20-fold more potent inhibitor of SNAV than Gal. A cDNA clone encoding SNAV was isolated from a cDNA library constructed with mRNA from the bark. Sequence analysis of this cDNA revealed a striking similarity to the recently cloned NeuAc a-2,6-gal/GalNAc-specific S. nigra bark agglutinin I (SNAI) and to the previously sequenced type-2 ribosome-inactivating proteins from Ricinus communis and Abrus precatorius. In addition, molecular modelling of SNAV further suggested that its structure closely resembles that of ricin. The Nterminal sequence of the B chain of SNAV also shows a marked similarity with the polypeptide of the previously described GalNAc-specific S. nigra bark agglutinin I1 (SNAII), which unlike SNAV and SNAI has no ribosome-inactivating activity. It appears, therefore, that elderbeny bark contains at least two different type-2 ribosome-inativating proteins and a lectin built up of subunits which are closely related to the B chain of SNAV.
Archives of Biochemistry and Biophysics, 1997
ribosomal RNA and stop protein synthesis (1). They A type II ribosome-inactivating protein (RIP) was are basically classified into two groups, i.e., type I (sinisolated from the bark tissue of Japanese elderberry gle chain) and type II (two chain) RIPs. Typical type II (Sambucus sieboldiana) and named sieboldin-b. Sie-RIPs such as ricin and abrin consist of a galactoseboldin-b is a heterodimeric protein consisting of 27specific lectin subunit, which serves to attach to the and 33-kDa subunits and showed strong ribosome-intarget cell surface by its ability to bind to galactoseactivating activity in vitro but did not show in vivo containing cell surface glycoconjugates, and a ribotoxicity. The amino acid sequence of sieboldin-b desome-inactivating N-glycosidase subunit (2). They are duced from the structure of the cDNA showed that highly toxic to mammalian cells and their possible both subunits of sieboldin-b are encoded on a single involvement in the plant defense system has been disprecursor polypeptide. Sieboldin-b has a structure hocussed (2). Recently, two novel RIPs which show strong mologous with the Neu5Ac(a2-6)Gal/GalNAc-specific ribosome-inactivating activity in vitro but do not show bark lectin from S. sieboldiana (SSA) and also typical in vivo toxicity were isolated from the leaves of Sambutype II RIPs such as ricin and abrin. Detailed analyses cus ebulus (ebulin-l) (3) and bark tissue of Sambucus of carbohydrate binding properties of sieboldin-b renigra (nigrin-b) (4).
Biochemical Journal, 2002
Sambucus nigra agglutinin I (SNA-I) is a type 2 ribosome-inactivating protein. Site-directed mutagenesis was used to mimic the conversion of the highly active B-chain of fruit-specific SNA (SNA-If) into the completely inactive B-chain of the closely related and naturally occurring loss-of-activity mutant called S. nigra agglutinin lectin-related protein. In the first mutant SNA-If-M1 the high-affinity site 2 of SNA-If was disrupted by replacing the presumed critical residue Asp231 with Glu231. In the double mutant SNA-If-M2, site 1 of SNA-If-M1 was also disrupted by substituting the presumed critical residue Asn48 with Ser48. The parent type 2 ribosome-inactivating protein and both mutants were expressed in Nicotiana tabacum Samsun NN and the recombinant proteins were purified and analysed. Recombinant SNA-If agglutinated rabbit erythrocytes equally well as SNA-If, but both mutants were completely inactive in this test. Binding assays to immobilized galactose and fetuin revealed tha...
FEBS Letters, 1998
Two novel highly basic type 1 (single chain) ribosomeinactivating proteins (RIPs) with N-glycosidase activity have been found in elderberries (the fruits of Sambucus nigra L.). Mass spectrometry of these RIPs, which we named nigritins f1 and f2, gave M r values of 24 095 and 23 565, respectively. Both proteins strongly inhibited protein synthesis in rabbit reticulocyte lysates but were inactive against plant ribosomes. Both nigritins have a similar topological activity on pBlueScript SK + DNA as that displayed by dianthin 30. Nigritin f1 is a constitutive RIP since it is present in both green and mature intact elderberries at nearly the same proportion with respect to total fruit protein. By contrast, nigritin f2 is inducible and only appeared in mature intact elderberries. Elderberries also contain two isoforms of a basic nigrin equivalent to the recently found basic nigrin b in elder bark (De Benito et al., FEBS Letters 413 (1997) 85^91). Our results indicate that probably not all plant RIPs exert the same biological function and that this may be determined by the physiological state of the tissue.
FEBS Letters, 1998
Two novel highly basic type 1 (single chain) ribosomeinactivating proteins (RIPs) with N-glycosidase activity have been found in elderberries (the fruits of Sambucus nigra L.). Mass spectrometry of these RIPs, which we named nigritins f1 and f2, gave M r values of 24 095 and 23 565, respectively. Both proteins strongly inhibited protein synthesis in rabbit reticulocyte lysates but were inactive against plant ribosomes. Both nigritins have a similar topological activity on pBlueScript SK + DNA as that displayed by dianthin 30. Nigritin f1 is a constitutive RIP since it is present in both green and mature intact elderberries at nearly the same proportion with respect to total fruit protein. By contrast, nigritin f2 is inducible and only appeared in mature intact elderberries. Elderberries also contain two isoforms of a basic nigrin equivalent to the recently found basic nigrin b in elder bark (De Benito et al., FEBS Letters 413 (1997) 85^91). Our results indicate that probably not all plant RIPs exert the same biological function and that this may be determined by the physiological state of the tissue. z 1998 Federation of European Biochemical Societies. Abbreviations: IC SH , concentration of RIP that gives 50% of inhibiton of translation in rabbit reticulocyte lysates; Nigritins f1 and f2, basic type 1 RIPs present in elderberry; rRNA, ribosomal ribonucleic acid; SDS-PAGE, polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate; RIP(s), ribosome-inactivating protein(s) FEBS 20262 FEBS Letters 428 (1998) 75^79
Journal of Biological Chemistry, 1997
One of the predominant proteins in the bark of elderberry (Sambucus nigra) has been identified as a novel type 2 ribosome-inactivating protein that exhibits a normal RNA N-glycosidase activity, but is devoid of carbohydrate binding activity. Sequence analysis of the corresponding cDNA clones revealed a striking homology to the previously cloned bark lectins from elderberry, suggesting that the new protein is a lectin-related protein. Molecular modeling of the protein confirmed that its A chain is fully active, whereas its B chain contains two functionally inactive carbohydrate-binding sites. These findings not only demonstrate for the first time the occurrence of a type 2 ribosome-inactivating protein with an inactive B chain, but also offer interesting perspectives for the synthesis of immunotoxins with an improved selectivity.
Journal of Experimental Botany, 1996
Sambucus (Caprifoliaceae) species contain nigrin b and ebulin I, which are two-chain ribosomeinactivating proteins (RIPs) belonging to a new type of RIPs which are non-toxic to mice and cultured human cells. In this work the presence in fruits of elder (S. nigra L.) of a new non-toxic type 2 RIP (nigrin f) that co-exists with a lectin known as SNA IV is described. Nigrin f strongly inhibited protein synthesis in mammalian, but not in plant, ribosomes, promoting the depurination of sensitive ribosomes and thus allowing the release of the RIP diagnostic RNA fragment. Nigrin f is composed of two dissimilar subunits linked by disulphide bridges with apparent M r values of 31 600 and 26 300. The N-terminal amino acid sequence revealed close homology of the catalytic A chain with type 1 RIPs, especially those from Cucurbitaceae, and the B chain with several lectins previously isolated from Sambucus species. Nigrin f was not toxic to mice when injected intraperitoneally up to 2mgkg~1. In addition, NHC human cells were also insensitive to nigrin f up to 60 //gml~1. Anti-nigrin b rabbit polyclonal antibodies reacted with nigrin f, indicating that nigrin b and nigrin f are proteins with similar structures.
Elderberries: A Source of Ribosome-Inactivating Proteins with Lectin Activity
Molecules, 2015
Sambucus (Adoxaceae) species have been used for both food and medicine purposes. Among these, Sambucus nigra L. (black elder), Sambucus ebulus L. (dwarf elder), and Sambucus sieboldiana L. are the most relevant species studied. Their use has been somewhat restricted due to the presence of bioactive proteins or/and low molecular weight compounds whose ingestion could trigger deleterious effects. Over the last few years, the chemical and pharmacological characteristics of Sambucus species have been investigated. Among the proteins present in Sambucus species both type 1, and type 2 ribosome-inactivating proteins (RIPs), and hololectins have been reported. The biological role played by these proteins remains unknown, although they are conjectured to be involved in defending plants against insect predators and viruses. These proteins might have an important impact on the nutritional characteristics and food safety of elderberries. Type 2 RIPs are able to interact with gut cells of insects and mammals triggering a number of