Identification and expression of NEU3, a novel human sialidase associated to the plasma membrane.
Several mammalian sialidases have been described so far, suggesting the existence of numerous polypeptides with different tissue distributions, subcellular localizations and substrate specificities. Among these enzymes, plasma-membrane-associated sialidase(s) have a pivotal role in modulating the ganglioside content of the lipid bilayer, suggesting their involvement in the complex mechanisms governing cell-surface biological functions. Here we describe the identification and expression of a human plasma-membrane-associated sialidase, NEU3, isolated starting from an expressed sequence tag (EST) clone. The cDNA for this sialidase encodes a 428-residue protein containing a putative transmembrane helix, a YRIP (single-letter amino acid codes) motif and three Asp boxes characteristic of sialidases. The polypeptide shows high sequence identity (78%) with the membrane-associated sialidase recently purified and cloned from Bos taurus. Northern blot analysis showed a wide pattern of expression of the gene, in both adult and fetal human tissues. Transient expression in COS7 cells permitted the detection of a sialidase activity with high activity towards ganglioside substrates at a pH optimum of 3.8. Immunofluorescence staining of the transfected COS7 cells demonstrated the protein's localization in the plasma membrane.
Monti
E
E
Telethon Institute of Genetics and Medicine (TIGEM), San Raffaele Biomedical Science Park, via Olgettina 58, 20132 Milan, Italy and Department of Biomedical Science and Biotechnology, University of Brescia, via Valsabbina 19, 25123 Brescia, Italy.
Bassi
M T
MT
Papini
N
N
Riboni
M
M
Manzoni
M
M
Venerando
B
B
Croci
G
G
Preti
A
A
Ballabio
A
A
Tettamanti
G
G
Borsani
G
G
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D019556:COS Cells; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D002880:Chromosomes, Human, Pair 11; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D020224:Expressed Sequence Tags; D005732:Gangliosides / Q000096:biosynthesis / Q000235:genetics; D015723:Gene Library; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009439:Neuraminidase / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D017422:Sequence Analysis, DNA; D013050:Spectrometry, Fluorescence; D014018:Tissue Distribution; D014162:Transfection
D018076:DNA, Complementary; D005732:Gangliosides; D008051:Lipid Bilayers; D009439:Neuraminidase