tailieunhanh - Báo cáo khoa học: Cloning and characterization of the genes encoding toxic lectins in mistletoe (Viscum album L)

Leaves of mistletoe (Viscum albumL)contain three toxic lectins (type 2 ribosome-inactivating proteins) MLI, MLII, and MLIII, differing in molecular mass and carbohydrate specificity. Clones, containing sequences of three gene vari-ants designatedml1p, ml2p,andml3p, were obtained using PCRamplification fromcDNAand frommistletoe genomic DNA. The quantitative ratio of theml1p, ml2p,andml3p genes in genomic DNA was found to be : 1 : 4, respectively, whereas the ratio of their mRNA was 50 : 10 : 1 | Eur. J. Biochem. 271 2350-2360 2004 FEBS 2004 doi Cloning and characterization of the genes encoding toxic lectins in mistletoe Viscum album L Alma G. Kourmanova1 2 Olga J. Soudarkina1 Sjur Olsnes3 and Jurij V. Kozlov1 2 1Engelhardt Institute of Molecular Biology Russian Academy of Sciences and 2The University of Oslo Centre for Medical Studies in Moscow Russia 3Institute for Cancer Research The Norwegian Radium Hospital Montebello Oslo Norway Leaves of mistletoe Viscum album L contain three toxic lectins type 2 ribosome-inactivating proteins MLI MLII and MLIII differing in molecular mass and carbohydrate specificity. Clones containing sequences of three gene variants designated ml1p ml2p and ml3p were obtained using PCR amplification from cDNA and from mistletoe genomic DNA. The quantitative ratio of the ml1p ml2p and ml3p genes in genomic DNA was found to be 1 4 respectively whereas the ratio of their mRNA was 50 10 1. The quantitative prevalence of the ml1p transcript correlates well with the observation that MLI is quantitatively dominant over MLII and MLIII in the mistletoe extract. The sequences of the proteins encoded by the ml1p ml2p and ml3p genes are identical to MLI by 98 88 and 77 respectively. The similarity to MLI of the amino acid sequence encoded by the gene ml1p the quantitative prevalent of its mRNA as well as structural properties of the B-chain indicate that the gene ml1p corresponds to MLI. Western blot analysis of recombinant A-chains encoded by the three variants of mlp genes with the monoclonal antibody MNA4 having differential affinity to MLI MLII and MLIII A-chains suggests that the ml2p and ml3p genes correspond to MLII and MLIII respectively. Structural differences in the carbohydrate-binding sites of the B-subunits of ML1p ML2p and ML3p probably explain the difference in sugar specificity of MLI MLII and MLIII. Keywords mistletoe ribosome-inactivating protein toxic lectin Viscum album viscumin. .

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