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R Léonard

Publications and source records attributed to R Léonard.

4 recordsLinked to original sources
PubMed2005

Biochemical characterization of Silene alba alpha4-fucosyltransferase and Lewis a products.

alpha1,4-Fucosylation has been recently detected in Arabidopsis thaliana [Leonard et al. (2002), Glycobiology 12: 299-306], and corresponding enzymes have also been characterized in Beta vulgaris [Bakker et al. (2001), FEBS Lett, 507: 307-312], and Lycopersicum aesculentum [Wilson (2001), Glycoconjugate J., 18: 439-447]. Here we demonstrated fucosyltransferase activity (FucT) in Silene alba cells and tissues. The Fuc linkage to GlcNAc residues of the lactosamine moiety of the Type I acceptor was confirmed by mass spectrometry experiments. Le(a)-glycoconjugates are found in the Golgi apparatus and plasma membrane of plant cells. In planta, the highest levels of activity were detected in seedlings, young roots and male flowers. The enzyme was stable up to 45( composite function)C and the optimum pH of reaction was 8.0. The enzyme required Mg(2+) or Mn(2+) for activity and was inhibited by Zn(2+) and ethylenediaminetetraacetic acid. Chemical modification of the enzyme with group-selective reagents revealed that selective modifications of arginine and lysine residues had no effect on enzyme activity. However the enzyme contains histidine and tryptophan residues that are essential for its activity. In contrast to human FUT3, the S. alba alpha4-FucT was insensitive to N-ethylmaleimide (NEM) treatment. Measurement of enzyme activity in S. alba cell fractions indicated that the enzyme is bound to microsomal membranes, furthermore a soluble isoform of the protein may be present.

Cell Membrane↗
PubMed2002

The presence of Lewis a epitopes in Arabidopsis thaliana glycoconjugates depends on an active alpha4-fucosyltransferase gene.

The presence of an alpha4-fucosyltransferase in plants was first deduced from the characterization of Lewis-a glycoepitopes in some N-glycans. The first plant gene encoding an alpha4-fucosyltransferase was recently cloned in Beta vulgaris. In the present paper we provide evidence for the presence of an alpha4-fucosyltransferase in A. thaliana by measurement of this glycosyltransferase activity from a purified microsomal preparation and by immunolocalization of Le(a) epitopes on glycans N-linked to glycoproteins located to the Golgi apparatus and on the cell surface. The corresponding gene AtFT4 (AY026941) was characterized. A unique copy of this gene was found in A. thaliana genome, and a single AtFT4 transcript was revealed in leaves, in roots, and at a lower extent in flowers. The coding sequence of AtFT4 gene is interrupted by two introns spanning 465 bp and 84 bp, respectively. The putative 393-amino-acid protein (44 kDa, pI: 6.59) contains an N-terminal hydrophobic region and one potential N-glycosylation site, but AtFT4 has poor homology (less than 30%) to the other alpha3/4-fucosyltransferases except for motif II. When expressed in COS 7 cells the protein is able to transfer Fuc from GDP-Fuc to a type 1 acceptor substrate, but this transferase activity is detected only in the culture medium of transfected cells

Amino Acid Sequence↗
PubMed1993

Modelling, synthesis and biological activity of a BLV proteinase, made of (only) 116 amino acids.

Bovine leukaemia virus (BLV) is the aetiological agent of Leukosis enzootica bovis [Viral Oncology (1980), G. Klein (Ed.) Raven Press, New York, pp. 231-238], a widely spread disease in cattle. BLV is reported as the animal model of human T-cell leukaemia virus (HLTV) which is the causative agent of adult T-cell leukaemia and tropical spastic paraparesis. Like the viruses themselves, the two retroviral proteinases (PR) are very closely related [Virology 142 (1985) 357-377]. BLV and HTLV-I PR are reported as putative proteins made of 126 [J. Virol. 57 (1986) 826-832] and 125 [FEBS Lett. 293 (1991) 106-110] amino acids, respectively (long sequences), belonging to the aspartyl proteinase family [Nature 329 (1987) 351-354], with the aid of molecular modelling, we show that BLV and HTLV-I proteinases made of only 116 and 115 amino acids, respectively (short sequences), display three-dimensional structures similar to that observed for other retroviral aspartyl proteinases. The models are based on three-dimensional structures of Rous sarcoma virus (RSV PR) and the human immunodeficiency virus (HIV-1 PR). We used solid phase peptide synthesis to produce the putative proteolytic enzyme of BLV (116 amino acids). In this study, we show that the folded synthetic protease accurately hydrolyzes a decapeptide corresponding to the sequence of the Matrice-Capside (MA/CA) cleavage site of the gag polyprotein. In addition, the proteolytic activity is inhibited by a statine ((4S,3S)-4-amino-3-hydroxyl-6-methylheptanoic acid) containing an analogous sequence.

Amino Acid Sequence↗