Primary hyperoxaluria type 1 in the Canary Islands: A conformational disease due to I244T mutation in the P11L-containing alanine:glyoxylate aminotransferase

Abstract

Primary hyperoxaluria type 1 (PH1) is an inborn error of metabo-lism resulting from a deficiency of alanine:glyoxylate aminotrans-ferase (AGXT; EC 2.6.1.44). Most of the PH1 alleles detected in theCanary Islands carry the Ile-2443Thr (I244T) mutation in theAGXTgene, with 14 of 16 patients homozygous for this mutation. Fourpolymorphisms withinAGXTand regional microsatellites also wereshared in their haplotypes (AGXT*LTM), consistent with a foundereffect. The consequences of these amino acid changes were inves-tigated. Although I244T alone did not affect AGXT activity orsubcellular localization, when present in the same protein moleculeas Leu-113Pro (L11P), it resulted in loss of enzymatic activity insoluble cell extracts. Like its normal counterpart, the AGXT*LTMprotein was present in the peroxisomes but it was insoluble indetergent-free buffers. The polymorphism L11P behaved as anintragenic modifier of the I244T mutation, with the resultingprotein undergoing stable interaction with molecular chaperonesand aggregation. This aggregation was temperature-sensitive.AGXT*LTM expressed inEscherichia coli, as a GST-fusion protein,and in insect cells could be purified and retained enzymatic activity.Among various chemical chaperones tested in cell culture, betainesubstantially improved the solubility of the mutant protein and theenzymatic activity in cell lysates. In summary, I244T, the secondmost common mutation responsible for PH1, is a protein confor-mational disease that may benefit from new therapies with phar-macological chaperones or small molecules to minimize protein aggregation.