Identification of a Sialidase Encoded in the Human Major Histocompatibility Complex

Abstract

Mammalian sialidases are important in modulating the sialic acid content of cell-surface and intracellular glycoproteins. However, the full extent of this enzyme family and the physical and biochemical properties of its individual members are unclear. We have identified a novel gene, G9, in the human major histocompatibility complex (MHC), that encodes a 415-amino acid protein sharing 21-28% sequence identity with the bacterial sialidases and containing three copies of the Asp-block motif characteristic of these enzymes. The level of sequence identity between human G9 and a cytosolic sialidase identified in rat and hamster (28 -29%) is much less than would be expected for analogous proteins in these species, suggesting that G9 is distinct from the cytosolic enzyme. Expression of G9 in insect cells has confirmed that it encodes a sialidase, which shows optimal activity at pH 4.6, but appears to have limited substrate specificity. The G9 protein carries an N-terminal signal sequence and immunofluorescence staining of COS7 cells expressing recombinant G9 shows localization of this sialidase exclusively to the endoplasmic reticulum. The location of the G9 gene, within the human MHC, corresponds to that of the murine Neu-1 locus, suggesting that these are analogous genes. One of the functions attributed to Neu-1 is the up-regulation of sialidase activity during T cell activation.

Molecular and structural studies of Japanese patients with sialidosis type 1

“…Recent cDNA cloning for human lysosomal neuraminidase has revealed that it encodes a 45-kDa protein with three potential N-linked glycosylation sites (Bonten et al 1996;Pshezhetsky et al 1997;Milner et al 1997). This information has facilitated the identification of gene mutations causing sialidosis.…”

“…According to the sequence alignment (Milner et al 1997), we built a model of the human lysosomal neuraminidase without the N-terminal 47 residues, corresponding to the signal peptide (Milner et al 1997;Vinogradova et al 1998). The model was, thus, composed of the 368 amino acids from residue 48 to the terminal residue 415, the amino acid sequence homology with the aligned fragment of Salmonella typhimurium LT2 being 39.7%.…”

“…The molecular basis of the disease has remained obscure. Recently, human lysosomal neuraminidase cDNA was isolated (Bonten et al 1996;Pshezhetsky et al 1997;Milner et al 1997), and studies for clarifying the pathogenesis of sialidosis were started. So far, six different mutations in the neuraminidase gene have been reported (Bonten et al 1996;Pshezhetsky et al 1997).…”

Molecular pathology of NEU1 gene in sialidosis

“…Cathepsin A was cloned and its mutations in galactosialidosis were first characterized 15 years ago [Galjart et al, 1988], but cloning of sialidase has been hampered for almost two decades by the low tissue content and instability of this enzyme. Three groups simultaneously identified the human sialidase cDNA and gene by a homologous search in the Expressed Sequence Tags Database (dbEST, National Center for Biotechnology Information) and direct sequencing of human chromosome 6 [Bonten et al, 1996;Milner et al, 1997;Pshezhetsky et al, 1997]. These studies paved the way for the characterization of the molecular basis of sialidosis.…”

Novel missense mutations in the human lysosomal sialidase gene in sialidosis patients and prediction of structural alterations of mutant enzymes

“…In recent years, the human lysosomal sialidase gene has been cloned (Bonten et al 1996;Pshezhetsky et al 1997;Milner et al 1997) on the basis of the genomic and structural information on the sialidases obtained from a variety of species, including viruses and bacteria (Roggentin et al 1989;Rothe et al 1991;Crennell et al 1993Crennell et al , 1994Gaskell et al 1995). The human sialidase cDNA contains a 1.2-kb open reading frame and a polyadenylation site within the 3Ј untranslated region.…”