Showing papers by "F. Van Leuven published in 1997"
TL;DR: These results demonstrate for the first time that a plant lectin is encoded by a truncated type 2 RIP gene but also address important questions with respect to the molecular evolution of RIP and lectins.
Abstract: The major protein of elderberry (Sambucus nigra L.) fruits is a lectin, called Sambucus nigra agglutinin IVf or SNAIVf. This lectin is composed of subunits that strongly resemble the B chain of the type 2 ribosome-inactivating protein (RIP), called SNAVf, present in the same tissue. To corroborate the possible relationship between both proteins their corresponding cDNAs were cloned and compared. Alignment of the deduced amino acid sequences revealed that the cDNA encoding SNAIVf is almost identical to that of SNAVf except that its A chain is truncated. Northern blot analysis confirmed that the mRNA encoding SNAIVf is about 500 nucleotides shorter than the SNAVf mRNA. In addition, the occurrence of a truncated type 2 RIP gene was unambiguously demonstrated by the analysis of PCR amplified genomic sequences. These results not only demonstrate for the first time that a plant lectin is encoded by a truncated type 2 RIP gene but also address important questions with respect to the molecular evolution of RIP and lectins.
47 citations
TL;DR: High-performance anion-exchange chromatography and gel-permeation analysis of the hydrolysis products indicated the preferential hydrolyzing of highly branched structures by the enzyme, suggesting that the enzyme action is similar to that by endoxylanases from Ceratocystis paradoxa, Aspergillus niger, and Neurospora crassa.
Abstract: An endogenous wheat (Triticum aestivum) flour endoxylanase was purified to homogeneity from a crude wheat flour extract by ammonium sulfate precipitation and cation-exchange chromatography The 30-kD protein had an isoelectric point of 93 or higher A sequence of 19 amino acids at the NH2 terminus showed 842% identity with an internal sequence of 15-kD grain-softness protein, friabilin High-performance anion-exchange chromatography and gel-permeation analysis of the hydrolysis products indicated the preferential hydrolysis of highly branched structures by the enzyme; wheat arabinoxylan and rye (Secale cereale) arabinoxylan (high arabinose to xylose ratios) were hydrolyzed more efficiently by this enzyme than oat (Avena sativa) spelt xylan (low arabinose to xylose ratios) The release of the hydrolysis products as a function of time suggested that the endoxylanolytic activity was associated with the release of arabinose units from the polysaccharides, suggesting that the enzyme action is similar to that by endoxylanases from Ceratocystis paradoxa, Aspergillus niger, and Neurospora crassa Although the enzyme released arabinose from arabinoxylan, it did not hydrolyze p-nitrophenyl-alpha-L-arabinofuranoside From the above, it follows that the enzyme, called arabinoxylanase, differs from most microbial endoxylanases and from an endoxylanase purified earlier from wheat flour
41 citations
TL;DR: Analysis of the lectin-alliinase complexes from garlic and ramsons bulbs demonstrated that only the heterodimeric lectins are capable of binding to the glycan chains of the alliinase molecules (EC 4.4.1.4).
Abstract: A procedure developed to separate the homodimeric and heterodimeric mannose-binding lectins from bulbs of garlic (Allium sativum L.) and ramsons (Allium ursinum L.) also enabled the isolation of stable lectin-alliinase complexes. Characterization of the individual lectins indicated that, in spite of their different molecular structure, the homomeric and heteromeric lectins resemble each other reasonably well with respect to their agglutination properties and carbohydrate-binding specificity. However, a detailed analysis of the lectin-alliinase complexes from garlic and ramsons bulbs demonstrated that only the heterodimeric lectins are capable of binding to the glycan chains of the alliinase molecules (EC 4.4.1.4). Moreover, it appears that only a subpopulation of the alliinase molecules is involved in the formation of lectin-alliinase complexes and that the complexed alliinase contains more glycan chains than the free enzyme. Finally, some arguments are given that the lectin-alliinase complexes do not occur in vivo but are formed in vitro after homogenization of the tissue.
35 citations
TL;DR: The accumulation of large quantities of type 1 RIP in a vegetative storage organ suggests that this presumed defence-related protein also plays a role in the nitrogen-storage metabolism of the bulb.
Abstract: The most abundant protein of Iris bulbs has been identified as a type 1 ribosome-inactivating protein (RIP). Analysis of the purified proteins and molecular cloning of the corresponding cDNAs demonstrated that this type 1 RIP is a mixture of three isoforms that exhibit a high degree of sequence identity and have similar, though not identical, ribosome-inactivating and polynucleotide:adenosine glycosidase activities. The accumulation of large quantities of type 1 RIP in a vegetative storage organ suggests that this presumed defence-related protein also plays a role in the nitrogen-storage metabolism of the bulb.
28 citations
TL;DR: Results suggest that (a) native-A2M binds to T. cruzi proteinase(s) and (b) T-cruzi expresses a receptor (s) that binds proteinase-complexed A2M.
Abstract: Trypanosoma cruzi, the causative agent of Chagas disease, infects vertebrate cells after an initial step of parasite/host-cell recognition. Alpha-2-macroglobulin (A2M), an important type of physiological proteinase inhibitor found in tissues and in the plasma of mammals, inhibits cell invasion by T. cruzi and accumulates in sites of the inflamed myocardium associated with parasite antigens. To study whether A2M would bind to T. cruzi, an indirect immunofluorescence reaction was performed using two different anti-mouse A2M sera. Intense labeling was observed in the membrane lining the cell body and the flagellum of bloodstream trypomastigotes obtained from experimentally infected mice in the peak of parasitemia, suggesting that the antisera recognize plasma A2M associated with the parasite surface. Metacyclic trypomastigotes obtained in a serum-free defined medium reacted with anti-A2M only after previous incubation with purified human A2M. Enzyme-linked immunosorbent assay (ELISA) studies were applied to characterize better the binding of native (N-A2M) and of proteinase-complexed (P-A2M) forms of A2M. The "in vitro" binding of N-A2M to trypomastigotes was better at pH 5.0, followed by pH 10.0 and pH 7.4. Cysteinly and serine proteinase inhibitors, E-64 and STI, respectively, inhibited the reaction. P-A2M also bound to T. cruzi in a dose-dependent way. Flow-cytometry studies showed that about 80% of the parasites stained with fluorescein isothiocyanate (FITC)-labeled P-A2M (50 micrograms/ml) with high affinity at pH 7.4 (but also at pH 10.0) in a process that was reverted by the addition of unlabeled P-A2M or the calcium-chelator agent EDTA and also by incubation at an acid pH (4.0). These results suggest that (a) native-A2M binds to T. cruzi proteinase(s) and (b) T. cruzi expresses a receptor(s) that binds proteinase-complexed A2M.
13 citations