PMID: 1533633

 

    Legend: Gene, Sites

Title : Analysis of the glycosylation and phosphorylation of the alpha- subunit of the lysosomal enzyme , beta-hexosaminidase A, by site-directed mutagenesis

Abstract :

1. The glycosylation and subsequent phosphorylation of mannose residues is a pivotal modification during the biosynthesis of lysosomal enzymes
2. We have identified the sites of N-linked glycosylation and oligosaccharide phosphorylation on the alpha- subunit of beta-hexosaminidase and have determined the influence of the oligosaccharides on the folding and transport of the enzyme
3. The potential glycosylation sequences , either singly or in combination, were eliminated through site-directed mutagenesis of the cDNA
4. By expression of the mutant cDNAs in COS-1 cells, each of the three glycosylation sites on the alpha- subunit was found to be modified by an oligosaccharide
5. One of the three oligosaccharides was the preferred site of phosphorylation
6. The absence of any individual oligosaccharide did not diminish the expression of the catalytic activity associated with the alpha-chain chain , implying proper folding and assembly of subunits
7. A profound effect was observed, however, when all three oligosaccharides were absent
8. The unglycosylated alpha- subunit , resulting from genetic alteration of all three glycosylation sites or synthesis of the wild-type protein in the presence of tunicamycin, was catalytically inactive
9. It was found to be improperly folded into an insoluble aggregate, linked through inappropriate disulfide bonds
10. The unglycosylated protein was trapped in the lumen of the endoplasmic reticulum and was found in a complex with the Ig heavy chain-binding protein , BiP
11. The properties of the nonglycosylated, misfolded alpha- subunit were similar to some mutant alpha- subunits in Tay-Sachs disease patients
12. The results indicate that the oligosaccharides are essential, although not in a site-specific manner, for proper folding and cellular transport of the alpha- subunit

Output (sent_index, trigger, protein, sugar, site):

• 0. glycosylation, , subunit, -, -
• 1. glycosylation, , -, mannose residues, -
• 10. unglycosylated, , protein, -, -
• 11. nonglycosylated, , subunit, -, -
• 2. glycosylation, , -, -, sites
• 4. glycosylation, , -, -, sites
• 4. modified, , -, an oligosaccharide, sites
• 8. unglycosylated, , subunit, -, -

Output(Part-Of) (sent_index, protein, site):

• 2. subunit, sites
• 4. subunit, sites

*Output_Site_Fusion* (sent_index, protein, sugar, site):

 

Protein	NCBI ID	SENTENCE INDEX
alpha-chain	2217	6