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Complete localization of disulfide bonds in GM2 activator protein

✍ Scribed by Christina G. Schütte; Thorsten Lemm; Konrad Sandhoff; Gereon J. Glombitza

Publisher
Cold Spring Harbor Laboratory Press
Year
2008
Tongue
English
Weight
628 KB
Volume
7
Category
Article
ISSN
0961-8368

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✦ Synopsis

Abstract

Lysosomal degradation of ganglioside GM2 by hexosaminidase A requires the presence of a small, non‐enzymatic cofactor, the GM2‐activator protein (GM2AP). Lack of functional protein leads to the AB variant of GM2‐gangliosidosis, a fatal lysosomal storage disease.

Although its possible mode of action and functional domains have been discussed frequently in the past, no structural information about GM2AP is available so far. Here, we determine the complete disulfide bond pattern of the protein. Two of the four disulfide bonds present in the protein were open to classical determination by enzymatic cleavage and mass spectrometry. The direct localization of the remaining two bonds was impeded by the close vicinity of cysteines 136 and 138. We determined the arrangement of these disulfide bonds by MALDI‐PSD analysis of disulfide linked peptides and by partial reduction, cyanylation and fragmentation in basic solution, as described recently (Wu F, Watson JT, 1997, Protein Sci 6:391‐398).

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