Requirement for small side chain residues within the GxGD-motif of presenilin for gamma-secretase substrate cleavage

J Neurochem. 2010 Feb;112(4):940-50. doi: 10.1111/j.1471-4159.2009.06510.x. Epub 2009 Dec 15.


gamma-Secretase is a pivotal intramembrane-cleaving protease complex and important drug target for Alzheimer's disease. The protease not only releases small peptides, such as the amyloid-beta peptide, which drives Alzheimer's disease pathogenesis, but also intracellular domains, which can have critical functions in nuclear signaling. Unlike typical aspartyl proteases, gamma-secretase contains a non-classical GxGD active site motif in its catalytic subunit presenilin (PS) 1 or PS2. It is not known whether both glycines are of similar functional relevance and why the glycine residues are invariant elements of the motif. Here we identify the N-terminal glycine of the GxGD motif in PS1, G382, as a critical residue of the active site domain of gamma-secretase. Substitution of G382 by a number of different amino acids abrogated gamma-secretase activity. Only the smallest possible G382A substitution allowed substantial gamma-secretase activity. Depending on the substrate, however, the presence of G382 could become even an absolute functional requirement of gamma-secretase. Very similar results were obtained for the C-terminal glycine residue (G384) of the GxGD motif. Our data thus identify a requirement for small side chain residues in the active site domain of gamma-secretase and suggest that the glycines of the GxGD motif could be evolutionary conserved to allow cleavage of all possible gamma-secretase substrates, including those, which are highly sensitive to minimal alteration of the PS active site domain. These findings broaden our understanding of gamma-secretase substrate recognition and cleavage, which may prove crucial for therapeutic targeting of the enzyme.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alanine / genetics
  • Amino Acid Motifs
  • Amyloid Precursor Protein Secretases / metabolism*
  • Amyloid beta-Peptides / metabolism
  • Amyloid beta-Protein Precursor / genetics
  • Amyloid beta-Protein Precursor / metabolism
  • Animals
  • Cells, Cultured
  • Embryo, Mammalian
  • Glycine / genetics*
  • Humans
  • Hyaluronan Receptors / genetics
  • Hyaluronan Receptors / metabolism
  • Immunoprecipitation
  • Mice
  • Mice, Knockout
  • Mutation / genetics
  • Presenilin-1 / chemistry*
  • Presenilin-1 / genetics
  • Presenilin-2 / chemistry*
  • Presenilin-2 / genetics
  • Protein Structure, Tertiary
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / methods
  • Transfection / methods


  • Amyloid beta-Peptides
  • Amyloid beta-Protein Precursor
  • Hyaluronan Receptors
  • Presenilin-1
  • Presenilin-2
  • Amyloid Precursor Protein Secretases
  • Alanine
  • Glycine