Molecular Insight into the Synergism between the Minor Allele of Human Liver Peroxisomal Alanine:Glyoxylate Aminotransferase and the F152I Mutation

J Biol Chem. 2009 Mar 27;284(13):8349-58. doi: 10.1074/jbc.M808965200. Epub 2009 Jan 20.

Abstract

Human liver peroxisomal alanine:glyoxylate aminotransferase (AGT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that converts glyoxylate into glycine. AGT deficiency causes primary hyperoxaluria type 1 (PH1), a rare autosomal recessive disorder, due to a marked increase in hepatic oxalate production. Normal human AGT exists as two polymorphic variants: the major (AGT-Ma) and the minor (AGT-Mi) allele. AGT-Mi causes the PH1 disease only when combined with some mutations. In this study, the molecular basis of the synergism between AGT-Mi and F152I mutation has been investigated through a detailed biochemical characterization of AGT-Mi and the Phe(152) variants combined either with the major (F152I-Ma, F152A-Ma) or the minor allele (F152I-Mi). Although these species show spectral features, kinetic parameters, and PLP binding affinity similar to those of AGT-Ma, the Phe(152) variants exhibit the following differences with respect to AGT-Ma and AGT-Mi: (i) pyridoxamine 5'-phosphate (PMP) is released during the overall transamination leading to the conversion into apoenzymes, and (ii) the PMP binding affinity is at least 200-1400-fold lower. Thus, Phe(152) is not an essential residue for transaminase activity, but plays a role in selectively stabilizing the AGT-PMP complex, by a proper orientation of Trp(108), as suggested by bioinformatic analysis. These data, together with the finding that apoF152I-Mi is the only species that at physiological temperature undergoes a time-dependent inactivation and concomitant aggregation, shed light on the molecular defects resulting from the association of the F152I mutation with AGT-Mi, and allow to speculate on the responsiveness to pyridoxine therapy of PH1 patients carrying this mutation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles*
  • Amino Acid Substitution
  • Genetic Diseases, Inborn / enzymology*
  • Genetic Diseases, Inborn / genetics
  • Glycine / genetics
  • Glycine / metabolism
  • Glyoxylates / metabolism
  • Humans
  • Hyperoxaluria / enzymology*
  • Hyperoxaluria / genetics
  • Liver / enzymology*
  • Mutation, Missense*
  • Oxalates / metabolism
  • Peroxisomes / enzymology*
  • Transaminases / deficiency
  • Transaminases / genetics
  • Transaminases / metabolism*

Substances

  • Glyoxylates
  • Oxalates
  • Transaminases
  • Alanine-glyoxylate transaminase
  • glyoxylic acid
  • Glycine