title: Molecular Insight into the Synergism between the Minor Allele of Human Liver Peroxisomal Alanine:Glyoxylate Aminotransferase and the F152I Mutation.
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.
authors.affiliation: Dipartimento di Scienze Morfologico-Biomediche, Sezione di Chimica Biologica, Facoltà di Medicina e Chirurgia, Università degli Studi di Verona, Strada Le Grazie, 8, 37134 Verona, Italy. carla.borrivoltattorni@univr.it
authors.forename: Barbara
authors.initials: B
authors.lastname: Cellini
authors.forename: Riccardo
authors.initials: R
authors.lastname: Montioli
authors.forename: Alessandro
authors.initials: A
authors.lastname: Paiardini
authors.forename: Antonio
authors.initials: A
authors.lastname: Lorenzetto
authors.forename: Carla Borri
authors.initials: CB
authors.lastname: Voltattorni
mesh_terms: D000483:Alleles; D019943:Amino Acid Substitution; D030342:Genetic Diseases, Inborn / Q000201:enzymology* / Q000235:genetics; D005998:Glycine / Q000235:genetics / Q000378:metabolism; D006038:Glyoxylates / Q000378:metabolism; D006801:Humans; D006959:Hyperoxaluria / Q000201:enzymology* / Q000235:genetics; D008099:Liver / Q000201:enzymology*; D020125:Mutation, Missense; D010070:Oxalates / Q000378:metabolism; D020675:Peroxisomes / Q000201:enzymology*; D000637:Transaminases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
chemical_list: D006038:Glyoxylates; D010070:Oxalates; D000637:Transaminases; C019635:Alanine-glyoxylate transaminase; C031150:glyoxylic acid; D005998:Glycine