Characterization of a Fungal Maleylacetoacetate Isomerase Gene and Identification of Its Human Homologue

Abstract

We have previously used Aspergillus nidulans as a fungal model for human phenylalanine catabolism. This model was crucial for our characterization of the human gene involved in alcaptonuria. We use here an identical approach to characterize at the cDNA level the human gene for maleylacetoacetate isomerase (MAAI, EC 5.2.1.2), the only as yet unidentified structural gene of the phenylalanine catabolic pathway.We report here the first characterization of a gene encoding a MAAI enzyme from any organism, the A. nidulans maiA gene. maiA disruption prevents growth on phenylalanine (Phe) and phenylacetate and results in the absence of MAAI activity in vitro and Phe toxicity. The MaiA protein shows strong amino acid sequence identity to glutathione S-transferases and has MAAI activity when expressed in Escherichia coli. maiA is clustered with fahA and hmgA, the genes encoding the two other enzymes of the common part of the Phe/phenylacetate pathways.Based on the high amino acid sequence conservation existing between other homologous A. nidulans and human enzymes of this pathway, we used the MaiA sequence in data base searches to identify human expressed sequence tags encoding its putative homologues. Four such cDNAs were sequenced and shown to be encoded by the same gene. They encode a protein with 45% sequence identity to MaiA, which showed MAAI activity when expressed in E. coli.Human MAAI deficiency would presumably cause tyrosinemia that would be characterized by the absence of succinylacetone, the diagnostic compound resulting from fumarylacetoacetate hydrolase deficiency in humans and fungi. Culture supernatants of an A. nidulans strain disrupted for maiA are succinylacetone-negative but specifically contain cis and/or trans isomers of 2,4-dioxohept-2-enoic acid. We suggest that this compound(s) might be diagnostic for human MAAI deficiency.

Molecular Evolution of Glutathione S-Transferases in the Genus Drosophila

“…The other z GST, CG9362, is evolving at a much faster rate. We therefore suggest that CG9363 is more likely to perform the maleylacetoacetate isomerase activity, highly conserved across eukaryotes, that functions in the catabolism of phenylalanine and tyrosine (Fernandez-Canon and Penalva 1998). CG9362 is likely to have a derived function, requiring less selective constraint.…”

Glutathione Transferases

“…GSTZs are highly conserved in eukaryotes, indicative of their important and central function in cell metabolism. The identifi cation of a fungal zeta GST as the remaining missing step in the catabolism of tyrosine led to the discovery of the enzyme's activity in catalysing the isomerisation of maleylacetoacetate to fumarylacetoacetate (Fernández-Cañón and Peñalva, 1998). GSTZ1 also effi ciently catalyses this reaction, and the presence of active upstream and downstream enzymes in the tyrosine catabolic pathway in Arabidopsis provides support that this pathway also operates in planta, even though technically it is not required Dixon and Edwards, 2006).…”

Immunohistochemical Localization and Activity of Glutathione Transferase Zeta (GSTZ1–1) in Rat Tissues

“…GSTZ1-1 is identical with maleylacetoacetate isomerase (Board et al, 1997;Fernández-Cañón and Peñalva, 1998), which catalyzes the penultimate step in tyrosine degradation (Knox and Edwards, 1955b). Hence, maleylacetoacetate is the endogenous substrate for GSTZ1-1 (Knox and Edwards, 1955a).…”