TY - JOUR
T1 - 4-Hydroxy-2-oxoglutarate aldolase inactivity in primary hyperoxaluria type 3 and glyoxylate reductase inhibition
AU - Riedel, Travis J.
AU - Knight, John
AU - Murray, Michael S.
AU - Milliner, Dawn S.
AU - Holmes, Ross P.
AU - Lowther, W. Todd
N1 - Funding Information:
We would like to thank Kendrah Kidd, Braden Gregory, Jennifer Steger, Lynnette C. Johnson, and Jill Clodfelter for their technical assistance. We would also like to thank Drs. R. Belostotsky, Y. Frishberg, and C. Monico for providing the identity of the HOGA PH3 mutations prior to their publication. This research was supported by grants from the Oxalosis and Hyperoxaluria Foundation and the National Institutes of Health ( DK083527 , DK054468 , DK073732 , and DK83908 ).
PY - 2012/10
Y1 - 2012/10
N2 - Mutations in the gene encoding for 4-hydroxy-2-oxoglutarate aldolase (HOGA) are associated with an excessive production of oxalate in Primary Hyperoxaluria type 3 (PH3). This enzyme is the final step of the hydroxyproline degradation pathway within the mitochondria and catalyzes the cleavage of 4-hydroxy-2-oxoglutarate (HOG) to pyruvate and glyoxylate. No analyses have been performed to assess the consequences of the mutations identified, particularly for those variants that produce either full-length or nearly full-length proteins. In this study, the expression, stability, and activity of nine PH3 human HOGA variants were examined. Using recombinant protein produced in Escherichia coli as well as transfected Chinese hamster ovary (CHO) cells, it was found that all nine PH3 variants are quite unstable, have a tendency to aggregate, and retain no measurable activity. A buildup of HOG was confirmed in the urine, sera and liver samples from PH3 patients. To determine how HOG is cleaved in the absence of HOGA activity, the ability of N-acetylneuraminate aldolase (NAL) to cleave HOG was evaluated. NAL showed minimal activity towards HOG. Whether the expected buildup of HOG in mitochondria could inhibit glyoxylate reductase (GR), the enzyme mutated in PH2, was also evaluated. GR was inhibited by HOG but not by 2-hydroxyglutarate or 2-oxoglutarate. Thus, one hypothetical component of the molecular basis for the excessive oxalate production in PH3 appears to be the inhibition of GR by HOG, resulting in a phenotype similar to PH2.
AB - Mutations in the gene encoding for 4-hydroxy-2-oxoglutarate aldolase (HOGA) are associated with an excessive production of oxalate in Primary Hyperoxaluria type 3 (PH3). This enzyme is the final step of the hydroxyproline degradation pathway within the mitochondria and catalyzes the cleavage of 4-hydroxy-2-oxoglutarate (HOG) to pyruvate and glyoxylate. No analyses have been performed to assess the consequences of the mutations identified, particularly for those variants that produce either full-length or nearly full-length proteins. In this study, the expression, stability, and activity of nine PH3 human HOGA variants were examined. Using recombinant protein produced in Escherichia coli as well as transfected Chinese hamster ovary (CHO) cells, it was found that all nine PH3 variants are quite unstable, have a tendency to aggregate, and retain no measurable activity. A buildup of HOG was confirmed in the urine, sera and liver samples from PH3 patients. To determine how HOG is cleaved in the absence of HOGA activity, the ability of N-acetylneuraminate aldolase (NAL) to cleave HOG was evaluated. NAL showed minimal activity towards HOG. Whether the expected buildup of HOG in mitochondria could inhibit glyoxylate reductase (GR), the enzyme mutated in PH2, was also evaluated. GR was inhibited by HOG but not by 2-hydroxyglutarate or 2-oxoglutarate. Thus, one hypothetical component of the molecular basis for the excessive oxalate production in PH3 appears to be the inhibition of GR by HOG, resulting in a phenotype similar to PH2.
KW - Calcium oxalate stone disease
KW - Hydroxyproline
KW - Oxalate
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U2 - 10.1016/j.bbadis.2012.06.014
DO - 10.1016/j.bbadis.2012.06.014
M3 - Article
C2 - 22771891
AN - SCOPUS:84864234719
SN - 0925-4439
VL - 1822
SP - 1544
EP - 1552
JO - Biochimica et Biophysica Acta - Molecular Basis of Disease
JF - Biochimica et Biophysica Acta - Molecular Basis of Disease
IS - 10
ER -