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Alkaptonuria (AKU; McKusick nº 203500) is a rare autosomal recessive disorder of both historical and medical interest. It represents a classical example of a discrete biochemical lesion resulting from a single gene deficiency that gives rise to a degenerative disease (O'Brien et al., 1963). AKU patients are deficient for homogentisate 1,2 dioxygenase activity (HGO, EC (La Du et al., 1958). This enzyme deficiency results in the accumulation of homogentisic acid (HGA), an intermediary metabolite in phenylalanine and tyrosine catabolism. In AKU patients, HGA is excreted in large amounts into the urine, which darkens on standing. This staining of the urine, which can be detected from early childhood, is the first and best known manisfestation of the disease and the one that brought it to clinical attention. Over the years, benzoquinone acetic acid (an oxidation product of HGA) is deposited either directly or as a polymer into connective tissues, causing their pigmentation (ochronosis) and eventually leading to serious arthropathy (Reviewed in La Du, 1995).

AKU was the first disease to be interpreted as a single gene trait. In 1902 Garrod reported the mode of inheritance in AKU (Garrod's original paper). He noted that affected individuals had normal parents and normal offspring and were frequently children of consanguineous marriages. Advised by Bateson, Garrod suggested that the peculiar form of heredity of AKU was best explained by the Mendelian theories of inheritance of a recessive character. In the 1908 Croonian Lectures to the Royal College of Physicians, Garrod unveiled the concept and the term inborn error of metabolism and went further to propose that AKU was caused by the lack of an enzyme that in normal individuals split the aromatic ring of HGA (Garrod, 1908). Biochemical evidence of the defect in AKU was provided by La Du in 1958, exactly fifty years later. He demonstrated the absence of HGO activity in a liver homogenate prepared from an AKU patient and established that the defect was limited to HGO, suggesting that in affected individuals there is a failure to synthesize active enzyme (La Du et al., 1958).

The gene responsible for AKU was located in man to 3q2 (Pollak et al.,1993 ; Janocha et al.,1994). In addition, a mutation causing alkaptonuria has been described in mouse and mapped to chromosome 16 (Montagutelli et al.,1994). Furthermore, a mouse homogentisate dioxygenase enzyme has been purified to homogeneity (Schmidt et al., 1995). However, the decisive contribution to the characterization of the human HGO gene came from work with the ascomycete fungus Aspergillus nidulans. Thus, a gene encoding an HGO enzyme in this organism, denominated hmgA, was cloned and characterised. The deduced amino acid sequence of its encoded protein product was used to identify EST (expressed sequence tag) clones putatively corresponding to the human HGO gene (Fernández-Cañón and Peñalva, 1995). With this ESTclone the human HGO gene was characterized and demonstrated that it is the AKU gene by showing that the AKU patients in two Spanish pedigrees were homozygous, or compound heterozygous, for the loss-of-function mutations P230S and V300G (Fernández-Cañón et al., 1996).

The human HGO gene is now completely sequenced (Granadino et al., 1997) and a remarkable number of AKU mutations have been already identified in patients from many different countries (Fernández-Cañón et al., 1996; Gehrig et al., 1997; Beltrán-Valero de Bernabé et al., 1998; Ramos et al., 1998; Higashino et al., 1998; Beltrán-Valero de Bernabé et al., 1999; Beltrán-Valero de Bernabé et al., 1999; Muller et al., 1999; Walter et al., 1999; Felbor et al., 1999;Porfirio et al., 2000; Zatkova et al., 2000a; unpublished data). Recently, the crystal structure of the human HGO enzyme has been determined (Titus et al., 2000), which has provided a framework for understanding the structural basis for the AKU mutations (Rodríguez et al., 2000).

The analysis and characterization of HGO intragenic polymorphisms has provided a general understanding of the variability at the HGO locus in both the AKU and the normal populations which has been very useful for the identification of AKU alleles and for tracing of their migration during recent human history (Beltrán-Valero de Bernabé et al., 1998). Analysis of the alkaptonuria mutations and polymorphisms has also revealed that the CCC sequence motif (or its reverse complement GGG) is a mutational hot spot in the HGO gene (Beltrán-Valero de Bernabé et al., 1999). Recent data suggest  that increased mutation rates in HGO associated to CCC triplets could have contributed to the remarkable high frequency of AKU found in particular geographical locations in Central Europe (Zatkova et al., 2000b).


Beltrán-Valero de Bernabé D., Granadino B., Chiarelli I., Porfirio B., Mayatepek E., Aquaron R., Moore M.M., Festen J.J., Sanmarti R., Peñalva M.A., Rodríguez de Córdoba S.R. (1998). Mutation and polymorphism analysis of the human homogentisate 1, 2-dioxygenase gene in alkaptonuria patients. Am. J. Hum. Genet., 62:776–784.

Beltrán-Valero de Bernabé D., Jimenez F.J., Aquaron R., Rodríguez de Córdoba S. (1999) Analysis of alkaptonuria (AKU) mutations and polymorphisms reveals that the CCC sequence motif is a mutational hot spot in the homogentisate 1,2 dioxygenase gene (HGO). Am. J. Hum. Genet., 64(5): 1316-22.

Beltrán-Valero de Bernabé D., Peterson P., Luopajarvi K., Matintalo P., Alho A., Konttinen Y., Krohn K., Rodríguez de Córdoba S., Ranki A. (1999) Mutational analysis of the HGO gene in Finnish alkaptonuria patients. J. Med. Genet., 36: 922-3.

Felbor U., Mutsch Y., Grehn F., Muller C.R., Kress W. (1999) Ocular ochronosis in alkaptonuria patients carrying mutations in the homogentisate 1,2-dioxygenase gene. Br. J. Ophthalmol., 83(6): 680-3.

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Fernández-Cañón J.M., Granadino B., Beltrán-Valero de Bernabé D., Renedo M., Fernández-Ruiz E., Peñalva M.A., Rodríguez de Córdoba S. (1996) The molecular basis of alkaptonuria. Nat. Genet., 14: 19–24.

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Janocha S., Wolz W., Srsen S., Srsnova K., Montagutelli X., Guenet J.L., Grimm T., Kress W., Muller C.R. (1994) The human gene for alkaptonuria (AKU) maps to chromosome 3q. Genomics, 19(1): 5-8.

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Muller C.R., Fregin A., Srsen S., Srsnova K., Halliger-Keller B., Felbor U., Seemanova E., Kress W. (1999) Allelic heterogeneity of alkaptonuria in Central Europe. Eur. J. Hum. Genet.,7(6): 645-51.

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Porfirio B., Chiarelli I., Graziano C., Mannoni A., Morrone A., Zammarchi E., Beltrán-Valero de Bernabé D., Rodríguez de Córdoba S. (2000). Alkaptonuria in Italy: polymorphic haplotype background, mutational profile, and description of four novel mutations in the homogentisate 1,2-dioxygenase gene. J. Med. Genet. 37:309-1

Ramos S.M., Hernandez M., Roces A., Larruga J.M., Gonzalez P., Gonzalez A.M., Pinto F.M., Cabrera V.M. (1998) Molecular diagnosis of alkaptonuria mutation by analysis of homogentisate 1,2 dioxygenase mRNA from urine and blood. Am. J. Med. Genet.,78(2):192-4.

Rodríguez JM, Timm DE, Titus GP, Beltrán-Valero de Bernabé D, Criado O, Mueller HA, Rodríguez de Córdoba S and Peñalva MA (2000) Structural and functional analysis of mutations in alkaptonuria. Hum Mol Genet 22:2341-2350.

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Titus G.P., Mueller H.A.A., Rodríguez de Córdoba S., Peñalva M.A., Timm D.E. (2000) Crystal structure of human homogentisate dioxygenase. Nat. Struct. Biol. 7(7):542-546.

Walter K., Gaa A., Schaefer H.E. (1999) Sequence analysis of the homogentisate 1,2 dioxygenase gene in a family affected by alkaptonuria. J. Med. Genet., 36(8): 645-6.

Zatkova A.,  Polakova H., Micutkova H.,  Zvarik M., Bosak V., Ferakova E., Matusek J., Ferak V., Kadasi L. (2000a). Novel mutations in homogentisate-1,2-dioxygenase gene identified in Slovak patients with alkaptonuria. J. Med. Genet. 37: 539-542.

Zatkova A., Beltrán-Valero de Bernabé D., Polakova H., Zvarik M., Ferakova E., Bosak V., Ferak V., Kadasi L., Rodríguez de Córdoba S. (2000b) High frequency of alkaptonuria in Slovakia. Evidence for the appearance of multiple mutations in HGO involving different mutational hot spots. Am. J. Hum. Genet., 67:1333-1339.