Homogentisate 1,2-dioxygenase Gene Research Articles

Homogentisate 1,2-dioxygenase (HGD) gene variants in young Egyptian patients with alkaptonuria

Alkaptonuria (AKU) is a rare autosomal recessive metabolic disorder caused by pathogenic variants in the homogentisate 1,2-dioxygenase (HGD) gene. This leads to a deficient HGD enzyme with the consequent accumulation of homogentisic acid (HGA) in different tissues causing complications in various organs, particularly in joints, heart valves and kidneys. The genetic basis of AKU in Egypt is completely unknown. We evaluated the clinical and genetic spectrum of six pediatric and adolescents AKU patients from four unrelated Egyptian families. All probands had a high level of HGA in urine by qualitative GC/MS before genetic confirmation by Sanger sequencing. Recruited AKU patients were four females and two males (median age 13 years). We identified four different pathogenic missense variants within HGD gene. Detected variants included a novel variant c.1079G > T;p.(Gly360Val) and three recurrent variants; c.1078G > C;p.(Gly360Arg), c.808G > A;p.(Gly270Arg) and c.473C > T;p.(Pro158Leu). All identified variants were properly segregating in the four families consistent with autosomal recessive inheritance. In this study, we reported the phenotypic and genotypic spectrum of alkaptonuria for the first time in Egypt. We further enriched the HGD-variant database with another novel pathogenic variant. The recent availability of nitisinone may promote the need for genetic confirmation at younger ages to start therapy earlier and prevent serious complications.

In-silico Profiling of Deleterious Non Synonymous SNPs of Homogentisate 1, 2 Dioxygenase (HGD) Gene for Early Diagnosis of “Alkaptonuria”

In-silico characterization and molecular modelling of a single amino acid substitution in HGD (Homogentisate 1,2dioxygenase) gene are mainly caused by the deficiency of enzyme Homogentisate 1,2dioxygenase (HGD). An enzyme HGD involved in the catabolism of amino acids such as tyrosine and phenylalanine. The objective of this study was to analyse non-synonymous SNPs from highly deleterious missense mutations which affect the protein function of HGD gene. Based on 3D structure different computational algorithms were performed to identify deleterious SNPs and assess the influence of mutation by using molecular dynamics simulations and molecular docking. Bioinformatics analysis like SIFT, PolyPhen 2.0, I mutant 3.0, PANTHER, SNPs and GO were performed to predict non deleterious ns-SNPs from missense mutations. Energy minimization was done by using GROMACS followed by RMSD calculations and free-energy values under SWISS-PDB viewer and PyMoL respectively. Later, Trajectory analysis was performed using computational tools like SRIDE, CONSURF, SPPIDER, PSIPRED, FLEXPRED for predicting the probably damaged ns-SNPs. Moreover, molecular docking was performed and identified highly deleterious probably damaging mutation. By operating 10 bioinformatics analysis, we obtained 5 mutations R53W, L61P, G121R, G361R and L430H which have an adverse effect on HGD gene. The results of the ConSurf analysis showed that all of these ns-SNPs are in the highly conserved positions and influence the structure of native proteins. L61P mutation had more effect on protein structure. Later, for future studies these mutations assists to develop an effective drug for the associated disease Alkaptonuria.

Ochronosis – a rare metabolic disease

Alkaptonuria is a rare disorder, an autosomal recessive condition with genetic determinism and hereditary transmission, having a prevalence of 1 per 1 million population in USA. The pathogenesis includes the deficiency of the homogentisate 1,2-dioxygenase (HGD) enzyme, an intermediary enzyme in phenylalanine and tyrosine catabolism. Mutations in HGD gene leads to deficient levels of functional HGD and an excess of homogentisic acid (HGA). Although HGA is rapidly excreted by the kidneys, it slowly accumulates in various tissues. Due to HGA oxidase deficiency, HGA turns into melanin-like pigment which determines: alkaptonuria, accumulation in the connective tissues, in the joints, or can make cardiovascular and genitourinary deposits. The chronic accumulation of HGA destroys the affected tissue, leading to the characteristic black-blue color and to clinical symptoms of alkaptonuria. The aim of this paper is to investigate the particularities of rheumatic manifestations in a rare metabolic disease and to support the correct diagnosis. A 58-year-old male patient was admitted to our clinic in 2019 for bilateral knee and left shoulder pain. In 2008 he was diagnosed with polyarticular ochronosis having dorsal and lumbar pain, mixed scapulohumeral pain, lumbar intervertebral disk calcifications; at that time, a diagnosis of ankylosing spondylitis or Forestier disease was excluded. At the current admission, the patient has been thoroughly reassessed to obtain a proper diagnosis and to determine the severity of the disease. The ochronotic axial damage caused important differential diagnosis problems with ankylosing spondylitis. Pigment deposition in the eyes, ears and skin does not cause problems to patients, but cardiovascular and genitourinary deposition leads to important complications. Kinetotherapy and NSAIDs are beneficial for pain symptoms. There is no specific medication for stopping the disease progression. Conclusions. Ochronosis is a rare disease which can cause a lot of problems regarding a proper diagnosis and treatment. When differential diagnosis with AS is difficult, the HLA-B27 genotyping is recommended. Final diagnosis is based on qualitative and quantitative urinary tests. The treatment includes only symptomatic drugs such as NSAIDs and kinetotherapy to improve joint mobility and muscle toning.

A molecular spectroscopy approach for the investigation of early phase ochronotic pigment development in Alkaptonuria

Alkaptonuria (AKU), a rare genetic disorder, is characterized by the accumulation of homogentisic acid (HGA) in organs due to a deficiency in functional levels of the enzyme homogentisate 1,2-dioxygenase (HGD), required for the breakdown of HGA, because of mutations in the HGD gene. Over time, HGA accumulation causes the formation of the ochronotic pigment, a dark deposit that leads to tissue degeneration and organ malfunction. Such behaviour can be observed also in vitro for HGA solutions or HGA-containing biofluids (e.g. urine from AKU patients) upon alkalinisation, although a comparison at the molecular level between the laboratory and the physiological conditions is lacking. Indeed, independently from the conditions, such process is usually explained with the formation of 1,4-benzoquinone acetic acid (BQA) as the product of HGA chemical oxidation, mostly based on structural similarity between HGA and hydroquinone that is known to be oxidized to the corresponding para-benzoquinone. To test such correlation, a comprehensive, comparative investigation on HGA and BQA chemical behaviours was carried out by a combined approach of spectroscopic techniques (UV spectrometry, Nuclear Magnetic Resonance, Electron Paramagnetic Resonance, Dynamic Light Scattering) under acid/base titration both in solution and in biofluids. New insights on the process leading from HGA to ochronotic pigment have been obtained, spotting out the central role of radical species as intermediates not reported so far. Such evidence opens the way for molecular investigation of HGA fate in cells and tissue aiming to find new targets for Alkaptonuria therapy.

Alkaptonuria in Turkey: Clinical and molecular characteristics of 66 patients

Alkaptonuria (AKU) is an inborn error of metabolism caused by the deficiency of homogentisate 1,2-dioxygenase (HGD) as a result of a defect in the HGD gene. HGD enzyme deficiency results in accumulation of homogentisic acid (HGA) in the body, which in turn leads to multisystemic clinical symptoms. The present study aimed to investigate the presenting symptoms, age at diagnosis, and clinical and genetic characteristics of AKU patients followed-up in different centers in Turkey. In this cross-sectional, multicenter, descriptive study, medical records of 66 AKU patients were retrospectively evaluated. Patients’ data regarding demographic, clinical and genetic characteristics were recorded. HGD database (http://hgddatabase.cvtisr.sk/) was used to identify HGD gene variants. Of the patients, 37 (56.1%) presented with isolated dark urine and 29 (43.9%) were diagnosed based on the clinical symptoms or family screening. One of these patients was on follow-up for 2 years due to Parkinsonism and was diagnosed with AKU on further analyses. Signs of ochronosis such as joint pain, low back pain and renal stones developed in childhood in 7 patients. Eight patients were diagnosed with depression via psychiatric evaluation. There were 14 (21.2%) patients operated on for ochronosis. The most frequent mutation observed in the patients was c.175delA, which was followed by c.674G > A and c.1007-2A > T mutations. Four novel mutations (c.189G > A, c.549+1G > T, c.1188+1G > A, and c.334 T > G) were identified in the patients included in the study. In addition to the known signs such as dark urine and skin pigmentation, symptoms involving different systems such as neurological findings and depression can also be encountered in AKU patients. The presence of a change in urine color needs to be questioned in patients presenting with different symptoms such as arthralgia/arthritis, renal stones or low-back pain, particularly in childhood, when skin ochronosis is not pronounced, and further examination should be performed.

A rare mutation in alkaptonuria patient

Alkaptonuria is a rare autosomal recessive metabolic disorder due to mutation in enzyme homogentisate 1,2-dioxygenase resulting in accumulation of homogentisic acid. The homogentisate 1,2-dioxygenase (HGD) gene has been mapped to chromosome 3q21- q23 and comprises 14 exons. A wide variety of causative mutations has been reported. Here, we are presenting a case report of a 2-year-old male child with a history of dark black-brown spots of urine on diaper with c.674G>C (p.Arg225Pro) mutation at exon 10 of HGD gene. The observed variant had a minor allele frequency of 0.0200% and 0.0004% in 1000 genomes and gnomAD database, respectively.

Founder effects of the homogentisate 1,2-dioxygenase (HGD) gene in a gypsy population and mutation spectrum in the gene among alkaptonuria patients from India.

Alkaptonuria (AKU) is a rare metabolic disease. The global incidence is 1:100,000 to 1:250,000. However, identification of a founder mutation in a gypsy population from India prompted us to study the prevalence of AKU in this population and to do molecular typing in referred cases of AKU from the rest of India. To determine the prevalence of AKU in the gypsy population predominantly residing in the seven districts of Tamil Nadu. To determine the molecular characteristic of AKU cases referred to our clinic from various parts of India. Urine spot test to detect homogentisic acid followed by quantitative estimation using high-performance liquid chromatography in 499 participants from the gypsy population and confirming the founder mutation in those with high levels by sequencing. Sequence thehomogentisate 1,2-dioxygenase (HGD) gene to identify mutations and variants in 29 AKU non-gypsy cases. Thefounder mutation was detected in homozygous state in 41/499 AKU-affected individuals of the gypsy community giving a high prevalence of 8.4%. Low back pain, knee pain, and eye and ear pigmentation were the most common symptoms and signs respectively. The commonest mutation identified in the non-gypsy AKU cases was p.Ala122Val. High prevalence of AKU in the inbred gypsy population at 8.4% was detected confirming the founder effect. Urine screening provided a cost-effective method to detect the disease early. Mutation spectrum is varied in the rest of the Indian population. This study identified maximum number of mutations in exon 6 of the HGD gene. Key Points • High prevalence (8.4%) of alkaptonuria (AKU) in the gypsy population due to founder mutation in the HGD gene. • Inbreeding exemplifies the founder effects of this rare genetic disorder. • Urinary screening is a cost-effective method in this community for early detection of AKU and intervention. • The mutation spectrum causing AKU is diverse in the rest of the Indian population.

Conditional targeting in mice reveals that hepatic homogentisate 1,2-dioxygenase activity is essential in reducing circulating homogentisic acid and for effective therapy in the genetic disease alkaptonuria.

Alkaptonuria is an inherited disease caused by homogentisate 1,2-dioxygenase (HGD) deficiency. Circulating homogentisic acid (HGA) is elevated and deposits in connective tissues as ochronotic pigment. In this study, we aimed to define developmental and adult HGD tissue expression and determine the location and amount of gene activity required to lower circulating HGA and rescue the alkaptonuria phenotype.We generated an alkaptonuria mouse model using a knockout-first design for the disruption of the HGD gene. Hgd tm1a −/− mice showed elevated HGA and ochronosis in adulthood. LacZ staining driven by the endogenous HGD promoter was localised to only liver parenchymal cells and kidney proximal tubules in adulthood, commencing at E12.5 and E15.5 respectively. Following removal of the gene trap cassette to obtain a normal mouse with a floxed 6th HGD exon, a double transgenic was then created with Mx1-Cre which conditionally deleted HGD in liver in a dose dependent manner. 20% of HGD mRNA remaining in liver did not rescue the disease, suggesting that we need more than 20% of liver HGD to correct the disease in gene therapy.Kidney HGD activity which remained intact reduced urinary HGA, most likely by increased absorption, but did not reduce plasma HGA nor did it prevent ochronosis. In addition, downstream metabolites of exogenous 13C6-HGA, were detected in heterozygous plasma, revealing that hepatocytes take up and metabolise HGA.This novel alkaptonuria mouse model demonstrated the importance of targeting liver for therapeutic intervention, supported by our observation that hepatocytes take up and metabolise HGA.

Twelve novel HGD gene variants identified in 99 alkaptonuria patients: focus on 'black bone disease' in Italy.

Alkaptonuria (AKU) is an autosomal recessive disorder caused by mutations in homogentisate-1,2-dioxygenase (HGD) gene leading to the deficiency of HGD enzyme activity. The DevelopAKUre project is underway to test nitisinone as a specific treatment to counteract this derangement of the phenylalanine-tyrosine catabolic pathway. We analysed DNA of 40 AKU patients enrolled for SONIA1, the first study in DevelopAKUre, and of 59 other AKU patients sent to our laboratory for molecular diagnostics. We identified 12 novel DNA variants: one was identified in patients from Brazil (c.557T>A), Slovakia (c.500C>T) and France (c.440T>C), three in patients from India (c.469+6T>C, c.650-85A>G, c.158G>A), and six in patients from Italy (c.742A>G, c.614G>A, c.1057A>C, c.752G>A, c.119A>C, c.926G>T). Thus, the total number of potential AKU-causing variants found in 380 patients reported in the HGD mutation database is now 129. Using mCSM and DUET, computational approaches based on the protein 3D structure, the novel missense variants are predicted to affect the activity of the enzyme by three mechanisms: decrease of stability of individual protomers, disruption of protomer-protomer interactions or modification of residues in the region of the active site. We also present an overview of AKU in Italy, where so far about 60 AKU cases are known and DNA analysis has been reported for 34 of them. In this rather small group, 26 different HGD variants affecting function were described, indicating rather high heterogeneity. Twelve of these variants seem to be specific for Italy.

Homogentisate 1,2 dioxygenase is expressed in brain: implications in alkaptonuria.

Alkaptonuria is an ultra-rare autosomal recessive disease developed from the lack of homogentisate 1,2-dioxygenase (HGD) activity, causing an accumulation in connective tissues of homogentisic acid (HGA) and its oxidized derivatives in polymerized form. The deposition of ochronotic pigment has been so far attributed to homogentisic acid produced by the liver, circulating in the blood, and accumulating locally. In the present paper, we report the expression of HGD in the brain. Mouse and human brain tissues were positively tested for HGD gene expression by western blotting. Furthermore, HGD expression was confirmed in human neuronal cells that also revealed the presence of six HGD molecular species. Moreover, once cultured in HGA excess, human neuronal cells produced ochronotic pigment and amyloid. Our findings indicate that alkaptonuric brain cells produce the ochronotic pigment in loco and this may contribute to induction of neurological complications.

한우 HGD 유전자내 변이지역과 경제형질간의 연관성 분석

The homogentisate 1,2-dioxygenase (HGD) gene, which consists of 14 exons and spans approximately 42630bp on Bos taurus autosome 1 (BTA 1), is one of the six enzymes required for catabolism of the aromatic amino acids tyrosine and phenylalanine. It has been reported that BTA1 harbors quantitative trait loci that effect marbling score (MS), carcass weight (CW), and longissimus muscle area (LMA) in cattle. The aim of this study was to identify the single nucleotide polymorphisms (SNPs) in the HGD gene and to analyze their association with economic traits in Korean cattle (Hanwoo). Genetic polymorphisms were screened by direct sequencing, which detected 10 SNPs (T11187C, T11301A, T11398G, G29833A, G34256T, G34257C, T34284C, T42333G, T42348C, and T42468C). Six polymorphic sites were selected for genotyping, and economic traits were analyzed using a general linear model in Korean cattle (n=90). The observed genotype frequencies for G34256T were 0.5843(GG), 0.3708(GT), and 0.0449(TT). In addition, 0.3596(GG), 0.3708(GC), and 0.2697(CC) were observed for the G34257C mutation. Statistical association analysis revealed that G34256T polymorphisms were significantly as-sociated with MS, and G34257C polymorphisms were significantly associated with MS and LMA (p<0.05). Further study is needed in order to use the genetic variant as a marker for marker-assisted selection in Korean cattle.

Mutation Screening of theHGDGene Identifies a Novel Alkaptonuria Mutation with Significant Founder Effect and High Prevalence

Alkaptonuria (AKU) is an autosomal recessive disorder; caused by the mutations in the homogentisate 1, 2-dioxygenase (HGD) gene located on Chromosome 3q13.33. AKU is a rare disorder with an incidence of 1: 250,000 to 1: 1,000,000, but Slovakia and the Dominican Republic have a relatively higher incidence of 1: 19,000. Our study focused on studying the frequency of AKU and identification of HGD gene mutations in nomads. HGD gene sequencing was used to identify the mutations in alkaptonurics. For the past four years, from subjects suspected to be clinically affected, we found 16 positive cases among a randomly selected cohort of 41 Indian nomads (Narikuravar) settled in the specific area of Tamil Nadu, India. HGD gene mutation analysis showed that 11 of these patients carry the same homozygous splicing mutation c.87 + 1G > A; in five cases, this mutation was found to be heterozygous, while the second AKU-causing mutation was not identified in these patients. This result indicates that the founder effect and high degree of consanguineous marriages have contributed to AKU among nomads. Eleven positive samples were homozygous for a novel mutation c.87 + 1G > A, that abolishes an intron 2 donor splice site and most likely causes skipping of exon 2. The prevalence of AKU observed earlier seems to be highly increased in people of nomadic origin.

Analysis of HGD Gene Mutations in Patients with Alkaptonuria from the United Kingdom: Identification of Novel Mutations.

Alkaptonuria (AKU) is a rare autosomal recessive disorder with incidence ranging from 1:100,000 to 1:250,000. The disorder is caused by a deficiency of the enzyme homogentisate 1,2-dioxygenase (HGD), which results from defects in the HGD gene. This enzyme converts homogentisic acid to maleylacetoacetate and has a major role in the catabolism of phenylalanine and tyrosine. To elucidate the mutation spectrum of the HGD gene in patients with alkaptonuria from 42 patients attending the National Alkaptonuria Centre, 14 exons of the HGD gene and the intron-exon boundaries were analysed by PCR-based sequencing. A total of 34 sequence variants was observed, confirming the genetic heterogeneity of AKU. Of these mutations, 26 were missense substitutions and four splice site mutations. There were two deletions and one duplication giving rise to frame shifts and one substitution abolishing the translation termination codon (no stop). Nine of the mutations were previously unreported novel variants. Using computational approaches based on the 3D structure, these novel mutations are predicted to affect the activity of the protein complex through destabilisation of the individual protomer structure or through disruption of protomer-protomer interactions.

Homogentisate 1,2 dioxygenase is expressed in human osteoarticular cells: Implications in alkaptonuria

Alkaptonuria (AKU) results from defective homogentisate1,2-dioxygenase (HGD), causing degenerative arthropathy. The deposition of ochronotic pigment in joints is so far attributed to homogentisic acid produced by the liver, circulating in the blood and accumulating locally. Human normal and AKU osteoarticular cells were tested for HGD gene expression by RT-PCR, mono- and 2D-Western blotting. HGD gene expression was revealed in chondrocytes, synoviocytes, osteoblasts. Furthermore, HGD expression was confirmed by Western blotting, that also revealed the presence of five enzymatic molecular species. Our findings indicate that AKU osteoarticular cells produce the ochronotic pigment in loco and this may strongly contribute to induction of ochronotic arthropathy. J. Cell. Physiol. 227: 3254–3257, 2012. © 2011 Wiley Periodicals, Inc.

Alkaptonuria in France: past experience and lessons for the future

Alkaptonuria (AKU) is an autosomal recessive disorder due to homogentisate 1,2-dioxygenase (HGD) deficiency in the liver and characterized by a triad of signs, according to chronology of appearance: homogentisic aciduria (HGA) or alkaptonuria, ochronosis then ochronotic arthropathy. This inborn error of metabolism is caused by mutations in the HGD gene. In this work we report observations of 96 AKU French patients from 81 families collected in the literature since 1882 and from our personal contribution since 1986, giving an incidence of the disease of around 1:680,000 (96/64.10(6)). As expected for an autosomal recessive disorder the main findings of this study were: a slight predominance of males (51/93, 54,8%) over females (42/93, 45,2%), a strong predominance of sibships with one affected individual (68/81, 84,0%) over sibships with two (11/81, 13.6%) and three(2/81, 2.4%) affected individuals. AKU families are scaterred among the French territory suggesting that most cases occured in non-consanguineous unions. Consanguinity was only found in five families. Other peculiarities of this study were (a) ten of these families have both parents from a foreign geographical origin: Poland(3), Italy(3), Portugal(2), Ukraine(1) and India(1) and four families with only one foreign parent (Algeria, Armenia, Serbia, UK), (b) HGD mutations were found in 23 families, (c) four of theses 96 patients were seen by us respectively 28, 29, 39 and 45 years after their report in the literature and (d) seven patients present cardiac and/or renal complications.

An update on molecular genetics of Alkaptonuria (AKU)

Alkaptonuria (AKU) is an autosomal recessive disorder caused by a deficiency of homogentisate 1,2 dioxygenase (HGD) and characterized by homogentisic aciduria, ochronosis, and ochronotic arthritis. The defect is caused by mutations in the HGD gene, which maps to the human chromosome 3q21-q23. AKU shows a very low prevalence (1:100,000-250,000) in most ethnic groups, but there are countries such as Slovakia and the Dominican Republic in which the incidence of this disorder rises to as much as 1:19,000. In this work, we summarize the genetic aspects of AKU in general and the distribution of all known disease-causing mutations reported so far. We focus on special features of AKU in Slovakia, which is one of the countries with an increased incidence of this rare metabolic disorder.

Meat quality and carcass traits in relation to HGD-BstXI and HGD-HaeIII PCR–RFLP polymorphism in Chinese red cattle

We investigated the effect of homogentisate 1, 2 dioxygenase (HGD) gene on meat quality and carcass traits in 287 Chinese red cattle. The PCR-SSCP method was used to identify polymorphism of the HGD gene in the exon 1 and intron 1. Two polymorphisms were detected in intron 1 and two restriction sites for endonuclease HGD-BstXI and HGD-HaeIII have also been found. The HGD-BstXI genotypes showed significant effects on cooking loss, drip loss, net meat weight, carcass weight, and eye muscle area (P<0.05). The HGD-HaeIII genotypes significant affected cooking loss, muscle fibre diameter, shear force, drip loss, and carcass yield ratio (P<0.05). Moreover, we found significant effects of diplotypes on cooking loss, muscle fibre diameter, shear force, drip loss, net meat weight, carcass weight, and eye muscle area (P<0.05).

Molecular cloning of the HGD gene and association of SNPs with meat quality traits in Chinese red cattle

Homogentisate 1, 2 dioxygenase (HGD) is one of six enzymes required for the catabolism of the aromatic amino acids phenylalanine and tyrosine. Here we present the nucleotide sequence of transcripts of the bovine HGD gene. The full length cDNA of bovine HGD has been identified, encoding a deduced protein of 445 amino acids (Accession No. FJ515744). The bovine HGD gene comprises 14 exons and 13 introns. This is the first published cDNA bovine sequences that share high sequence similarity with other species. Semi-quantitative RT-PCR analysis demonstrated that the bovine HGD transcript was mainly expressed in liver and kidney tissues. Nine single nucleotide polymorphisms (SNPs) were identified, five in the coding region and four intronic. Four of the SNPs change an amino acid in the HGD protein sequence. Genotype and allelic frequencies were determined in Chinese red cattle breeds. Ten haplotypes were determined based upon the genotype of 9 SNPs. Moreover, for the first time an association was reported between HGD gene polymorphism and meat quality traits in Chinese red cattle (n = 224). Marker-trait association analyses showed that the HGD/PvuII genotypes showed a significant effect on meat cooking rate, muscle fiber diameter, and shear force (P < 0.05). The HGD DraIII genotypes showed a significant effect on muscle fiber diameter, shear force, and drip loss (P < 0.05). The HGD/AluI genotypes showed a significant effect on meat cooking rate, shear force, and drip loss (P < 0.05). The HGD/DraI genotypes showed a significant effect on meat cooking rate and shear force (P < 0.05). The HGD/EcoRV genotypes showed a significant effect on meat cooking rate, muscle fiber diameter, and shear force (P < 0.05). In all loci, no statistically significant differences were observed for pHu (P > 0.05). This is the first incidence where polymorphisms of a bovine HGD gene have demonstrated a correlation with meat traits in Chinese red cattle.

A novel missense HGD gene mutation, K57N, in a patient with alkaptonuria

Alkaptonuria is a rare recessive disorder of phenylalanine/tyrosine metabolism due to a defect in the enzyme homogentisate 1,2-dioxygenase (HGD) caused by mutations in the HGD gene. We report the case of a 38 year-old male with known alkaptonuria who was referred to an adult metabolic clinic after initially presenting to an emergency department with renal colic and subsequently passing black ureteric calculi. He complained of severe debilitating lower back pain, worsening over the last few years. A CT scan revealed marked degenerative changes and severe narrowing of the disc spaces along the entire lumbar spine. Sequencing of the HGD gene revealed that he was a compound heterozygote for a previously described missense mutation in exon 13 (G360R) and a novel missense mutation in exon 3 (K57N). Lys 57 is conserved among species and mutation of this residue is predicted to affect HGD protein function by interfering with substrate traffic at the active site. In summary, we describe an alkaptonuric patient and report a novel missense HGD mutation, K57N.

Three‐generational alkaptonuria in a non‐consanguineous family

Alkaptonuria (AKU) is a rare inborn error of metabolism of aromatic amino acids and considered to be an autosomal recessive trait caused by mutations in the homogentisate 1,2-dioxygenase (HGD) gene. A dominant pattern of inheritance has been reported but was attributed to extended consanguinity in many cases. However, we have observed a non-consanguineous family segregating AKU in a dominant manner over three generations. All affected individuals presented with typical features of AKU including darkening of the urine, ochronosis, arthropathy, and elevated urinary excretion of homogentisic acid. Sequence analysis of the HGD gene from genomic DNA of two affected individuals, uncle and niece, revealed a heterozygous missense mutation (M368V) in the uncle that was not present in his niece. Microsatellite genotyping demonstrated that both were heterozygous at the HGD locus and shared one haplotype. This haplotype did not contain a detectable HGD mutation. The haplotype was also found in a healthy son of the niece, making a dominant HGD mutation unlikely. Moreover, sequencing of cDNA from lymphoblastoid cells of the niece did not reveal an HGD mRNA with a potentially dominant-negative effect. Rare causes of the uncommon AKU inheritance in this family have to be considered, ranging from the coincidence of undetectable HGD mutations to a dominant mutation of a second, hitherto unknown AKU gene.