Reexamination of aspartoacylase: Is this human enzyme really a glycoprotein?

• Defects in the enzyme aspartoacylase lead to a fatal neurological disease.
• Our previous work suggested the presence of an essential N-glycosylation.
• More extensive studies have failed to detect the presence of an glycan in this enzyme.
• Alternative interpretations are presented for these previous studies.
• Definitive evidence is presented to show that the recombinantly-expressed human enzyme is not a glycoprotein.

Aspartoacylase catalyzes the metabolism of an important amino acid in the brain, with the release acetate serving as the source for fatty acid biosynthesis. Defects in this enzyme lead to a loss of activity and the symptoms of a fatal neurological disorder called Canavan disease. Extensive evidence, including deglycosylation studies, differential activity upon eukaryotic host expression and site directed mutagenesis, have supported the presence of a glycan that plays an essential role in the stability and catalytic activity of mammalian aspartoacylase. However, the structure of this enzyme did not show the presence of any non-amino acid components at the putative glycosylation site. A more extensive study specifically designed to resolve this discrepancy has now shown that recombinantly-expressed human aspartoacylase is not glycosylated, but is still fully functional and stable even when produced from a bacterial expression system. Alternative interpretations of the prior experiments now present a consistent picture of the structural components of this essential brain enzyme.