PMID: 11320060Apr 26, 2001Paper

The DXD motif is required for GM2 synthase activity but is not critical for nucleotide binding

Glycobiology
J LiW W Young

Abstract

We tested the importance of the aspartate-any residue-aspartate (DXD) motif for the enzymatic activity and nucleotide binding capacity of the Golgi glycosyltransferase GM2 synthase. We prepared point mutations of the motif, which is found in the sequence 352-VLWVDDDFV, and analyzed cells that stably expressed the mutated proteins. Whereas the folding of the mutated proteins was not seriously disrupted as judged by assembly into homodimers, Golgi localization, and secretion of a soluble form of the enzyme, exchange of the highly conserved aspartic acid residues at position 356 or 358 with alanine or asparagine reduced enzyme activity to background levels. In contrast, the D356E and D357N mutations retained weak activity, while the activity of V352A and W354A mutants was 167% and 24% that of wild-type enzyme, respectively. Despite the major effect of the DXD motif on enzymatic activity, nucleotide binding was not altered in the triple mutant D356N/D357N/D358N as revealed by binding to UDP-beads and labeling with the photoaffinity reagent, P(3)-(4-azidoanilido)uridine 5'-triphosphate (AAUTP). In summary, rather than being critical for nucleotide binding, this motif may function during catalysis in GM2 synthase, as has been propose...Continue Reading

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Oct 6, 2000·The Journal of Biological Chemistry·J LiW W Young

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Citations

Nov 6, 2002·Biochimica Et Biophysica Acta·Beverly M ZakJeffrey D Esko
Jun 15, 2011·The Journal of Biological Chemistry·Hiromu TakematsuYasunori Kozutsumi
May 28, 2009·Glycobiology·Jennifer T AguilanPamela Stanley
Oct 18, 2015·Cellular and Molecular Life Sciences : CMLS·Sakari KellokumpuTuomo Glumoff
Mar 23, 2012·The Journal of Physical Chemistry. B·Laura DuitchReinhard Schweitzer-Stenner
Aug 6, 2004·The Journal of Biological Chemistry·Christian GöttingKnut Kleesiek
May 25, 2007·The Journal of Biological Chemistry·Shao-Yang KuP Lynne Howell
Jan 30, 2007·The Journal of Biological Chemistry·Mattias PerssonMartin L Olsson
Jun 10, 2003·The Journal of Biological Chemistry·Sandrine GulbertiMohamed Ouzzine

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