In normal human hepatocytes, the intermediary-metabolic enzyme alanine:glyoxylate aminotransferase (AGT) is located within the peroxisomes. However, in approx. one-third of patients suffering from the hereditary kidney stone disease primary hyperoxaluria type 1, AGT is mistargeted to the mitochondria. AGT mistargeting results from the synergistic interaction between a common P11L (Pro11→Leu) polymorphism and a disease-specific G170R mutation. The polymorphism generates a functionally weak mitochondrial targeting sequence, the efficiency of which is increased by the mutation. The two substitutions together, but not in isolation, inhibit AGT dimerization, highlighting the different structural requirements of the peroxisomal and mitochondrial protein-import machineries. In the present study, we show that treatments known to increase the stability of proteins non-specifically (i.e. lowering the temperature from 37 to 30 °C or by the addition of glycerol) completely normalize the intracellular targeting of mutant AGT expressed in transfected COS cells. On the other hand, treatments known to decrease protein stability (e.g. increasing the temperature from 37 to 42 °C) exacerbate the targeting defect. Neither of the treatments affects the relative efficiencies of the peroxisomal and mitochondrial protein-import pathways intrinsically. Results are discussed in the light of the known structural requirements of the two protein trafficking pathways and the formulation of possible treatment strategies for primary hyperoxaluria type 1.
- alanine:glyoxylate aminotransferase
- enzyme trafficking defect
- kidney stone disease
- mitochondrial protein targeting
- peroxisomal protein targeting
- primary hyperoxaluria type 1
Abbreviations used: AGT, alanine:glyoxylate aminotransferase; CFTR, cystic fibrosis transmembrane conductance regulator; GFP, green fluorescent protein; MTS, mitochondrial targeting sequence; PH1, primary hyperoxaluria type 1; PTS1, peroxisomal targeting sequence type 1; for brevity the one-letter system for amino acids has been used: P11L, for example, means Pro11→Leu.
- The Biochemical Society, London ©2003