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The Chaperoning Activity of Amino-oxyacetic Acid on Folding-Defective Variants of Human Alanine:Glyoxylate Aminotransferase Causing Primary Hyperoxaluria Type I.

Abstract
The rare disease Primary Hyperoxaluria Type I (PH1) results from the deficit of liver peroxisomal alanine:glyoxylate aminotransferase (AGT), as a consequence of inherited mutations on the AGXT gene frequently leading to protein misfolding. Pharmacological chaperone (PC) therapy is a newly developed approach for misfolding diseases based on the use of small molecule ligands able to promote the correct folding of a mutant enzyme. In this report, we describe the interaction of amino-oxyacetic acid (AOA) with the recombinant purified form of two polymorphic species of AGT, AGT-Ma and AGT-Mi, and with three pathogenic variants bearing previously identified folding defects: G41R-Ma, G170R-Mi, and I244T-Mi. We found that for all these enzyme AOA (i) forms an oxime at the active site, (ii) behaves as a slow, tight-binding inhibitor with KI values in the nanomolar range, and (iii) increases the thermal stability. Furthermore, experiments performed in mammalian cells revealed that AOA acts as a PC by partly preventing the intracellular aggregation of G41R-Ma and by promoting the correct peroxisomal import of G170R-Mi and I244T-Mi. Based on these data, we carried out a small-scale screening campaign. We identified four AOA analogues acting as AGT inhibitors, even if only one was found to act as a PC. The possible relationship between the structure and the PC activity of these compounds is discussed. Altogether, these results provide the proof-of-principle for the feasibility of a therapy with PCs for PH1-causing variants bearing folding defects and provide the scaffold for the identification of more specific ligands.
AuthorsElisa Oppici, Riccardo Montioli, Mirco Dindo, Laura Maccari, Valentina Porcari, Antonio Lorenzetto, Sara Chellini, Carla Borri Voltattorni, Barbara Cellini
JournalACS chemical biology (ACS Chem Biol) Vol. 10 Issue 10 Pg. 2227-36 (Oct 16 2015) ISSN: 1554-8937 [Electronic] United States
PMID26161999 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References
  • Molecular Chaperones
  • Aminooxyacetic Acid
  • Transaminases
  • glyoxylate aminotransferase
  • Alanine
Topics
  • Alanine (genetics)
  • Aminooxyacetic Acid (chemistry, metabolism, pharmacology)
  • Blotting, Western
  • Fluorescent Antibody Technique
  • Genetic Variation
  • Humans
  • Hyperoxaluria, Primary (enzymology, genetics)
  • Molecular Chaperones (metabolism)
  • Protein Folding (drug effects)
  • Protein Stability
  • Transaminases (genetics, metabolism)

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