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The Biochemical Journal Sep 1992Aspartylglucosaminidase (AGA, EC 3.5.1.26) is an essential enzyme in the degradation of asparagine-linked glycoproteins. In man, deficient activity of this enzyme leads...
Aspartylglucosaminidase (AGA, EC 3.5.1.26) is an essential enzyme in the degradation of asparagine-linked glycoproteins. In man, deficient activity of this enzyme leads to aspartylglucosaminuria (AGU), a recessively inherited lysosomal storage disease. Here we used affinity-purified polyclonal antibodies against the native AGA and its denatured subunits to establish the molecular structure and intracellular location of the enzyme in normal and AGU fibroblasts. Inactivation of the enzyme was found to coincide with the dissociation of the heterodimeric enzyme complex into subunits. Although the subunits were not linked by covalent forces, the intrapolypeptide disulphide bridges were found to be essential for the normal function of AGA. AGA was localized into lysosomes in control fibroblasts by both immunofluorescence microscopy and immuno-electron microscopy, whereas in AGU cells the location of antigen was different, suggesting that, owing to the mutation, a missing disulphide bridge, most of the enzyme molecules get retarded in the cis-Golgi region and most probably face intracellular degradation.
Topics: Aspartylglucosaminuria; Aspartylglucosylaminase; Blotting, Western; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Fibroblasts; Humans; Immunohistochemistry; Leukocytes; Microscopy, Fluorescence; Microscopy, Immunoelectron; Peptide Fragments; Precipitin Tests; Temperature
PubMed: 1530592
DOI: 10.1042/bj2860613 -
The Biochemical Journal Mar 2004Aspartylglucosaminidase (AGA) belongs to the N-terminal nucleophile (Ntn) hydrolase superfamily characterized by an N-terminal nucleophile as the catalytic residue....
Aspartylglucosaminidase (AGA) belongs to the N-terminal nucleophile (Ntn) hydrolase superfamily characterized by an N-terminal nucleophile as the catalytic residue. Three-dimensional structures of the Ntn hydrolases reveal a common folding pattern and equivalent stereochemistry at the active site. The activation of the precursor polypeptide occurs autocatalytically, and for some amidohydrolases of prokaryotes, the precursor structure is known and activation mechanisms are suggested. In humans, the deficient AGA activity results in a lysosomal storage disease, aspartylglucosaminuria (AGU) resulting in progressive neurodegeneration. Most of the disease-causing mutations lead to defective molecular maturation of AGA, and, to understand the structure-function relationship better, in the present study, we have analysed the effects of targeted amino acid substitutions on the activation process of human AGA. We have evaluated the effect of the previously published mutations and, in addition, nine novel mutations were generated. We could identify one novel amino acid, Gly258, with an important structural role on the autocatalytic activation of human AGA, and present the molecular mechanism for the autoproteolytic activation of the eukaryotic enzyme. Based on the results of the present study, and by comparing the available information on the activation of the Ntn-hydrolases, the autocatalytic processes of the prokaryotic and eukaryotic enzymes share common features. First, the critical nucleophile functions both as the catalytic and autocatalytic residue; secondly, the side chain of this nucleophile is oriented towards the scissile peptide bond; thirdly, conformational strain exists in the precursor at the cleavage site; finally, water molecules are utilized in the activation process.
Topics: Amino Acids; Animals; Aspartylglucosylaminase; COS Cells; Catalysis; Chlorocebus aethiops; Enzyme Activation; Humans; Models, Molecular; Mutagenesis, Site-Directed; Threonine
PubMed: 14616088
DOI: 10.1042/BJ20031496 -
The American Journal of Pathology Oct 1998Aspartylglycosaminuria (AGU) is one of the most common lysosomal storage disorders in humans. A mouse model for AGU has been recently generated through targeted...
Aspartylglycosaminuria (AGU) is one of the most common lysosomal storage disorders in humans. A mouse model for AGU has been recently generated through targeted disruption of the glycosylasparaginase gene, and at a young age the glycosyl asparaginase-deficient mice demonstrated many pathological changes found in human AGU patients (Kaartinen V, Mononen I, Voncken J-W, Gonzalez-Gomez I, Heisterkamp N, Groffen J: A mouse model for aspartylglycosaminuria. Nat Med 1996, 2:1375-1378). Our current findings demonstrate that after the age of 10 months, the general condition of null mutant mice gradually deteriorated. They suffered from a progressive motoric impairment and impaired bladder function and died prematurely. A widespread lysosomal hypertrophy in the central nervous system was detected. This neuronal vacuolation was particularly severe in the lateral thalamic nuclei, medullary reticular nuclei, vestibular nuclei, inferior olivary complex, and deep cerebellar nuclei. The oldest animals (20 months old) displayed a clear neuronal loss and gliosis, particularly in those regions, where the most severe vacuolation was found. The severe ataxic gait of the older mice was likely due to the dramatic loss of Purkinje cells, intensive astrogliosis and vacuolation of neurons in the deep cerebellar nuclei, and the severe vacuolation of the cells in vestibular and cochlear nuclei. The impaired bladder function and subsequent hydronephrosis were secondary to involvement of the central nervous system. These findings demonstrate that the glycosylasparaginase-deficient mice share many neuropathological features with human AGU patients, providing a suitable animal model to test therapeutic strategies in the treatment of the central nervous system effects in AGU.
Topics: Acetylglucosamine; Animals; Aspartylglucosaminuria; Aspartylglucosylaminase; Central Nervous System; Cytoplasm; Female; Gait; Glial Fibrillary Acidic Protein; Immunoenzyme Techniques; Lysosomal Storage Diseases; Lysosomes; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neurodegenerative Diseases; Neurons; Urinary Bladder; Vacuoles
PubMed: 9777961
DOI: 10.1016/S0002-9440(10)65674-X -
Neurology. Genetics Jun 2024The causes of intellectual disability (ID) are varied, with as many as 1,400 causative genes. We attempted to identify the causative gene in a patient with long-standing...
OBJECTIVES
The causes of intellectual disability (ID) are varied, with as many as 1,400 causative genes. We attempted to identify the causative gene in a patient with long-standing undiagnosed ID.
METHODS
Although this was an isolated case with no family history, we searched for the causative gene using trio-based whole-exome sequencing (trio-WES), because severe ID is often caused by genetic variations, and inherited metabolic disorders (IMDs) are assumed to be the cause when regression and epilepsy occur.
RESULTS
We identified homozygous donor splice-site variants in the gene (aspartylglucosaminidase; NM_000027.4) Chr4(GRCh38):g. 177436275C>A, c.698+1G>T. This gene is implicated in aspartylglucosaminuria (AGU; OMIM #208400) and originated from both of the patient's parents. We confirmed the pathogenicity of the variant by detecting the splicing defect in cDNA from the patient's blood and accumulation of aberrant metabolites in the patient's urine.
DISCUSSION
We discuss how to more readily achieve an accurate diagnosis for patients with undiagnosed intellectual disabilities. Medical practitioners' awareness of the characteristics of the disease leading to clinical suspicion in patients with matching presentations, and the performance of newborn screening when possible, is important for the diagnosis of ID. In addition, the characteristic symptoms and course of the disease give rise to suspicion of IMDs. Given our results, we consider trio-WES to be a powerful method for identifying the causative genes in cases of ID with genetic causes.
PubMed: 38831911
DOI: 10.1212/NXG.0000000000200161 -
The EMBO Journal Jan 1991We have isolated a 2.1 kb cDNA which encodes human aspartylglucosaminidase (AGA, E.C. 3.5.1.26). The activity of this lysosomal enzyme is deficient in...
We have isolated a 2.1 kb cDNA which encodes human aspartylglucosaminidase (AGA, E.C. 3.5.1.26). The activity of this lysosomal enzyme is deficient in aspartylglucosaminuria (AGU), a recessively inherited lysosomal accumulation disease resulting in severe mental retardation. The polypeptide chain deduced from the AGA cDNA consists of 346 amino acids, has two potential N-glycosylation sites and 11 cysteine residues. Transient expression of this cDNA in COS-1 cells resulted in increased expression of immunoprecipitable AGA protein. Direct sequencing of amplified AGA cDNA from an AGU patient revealed a G----C transition resulting in the substitution of cysteine 163 with serine. This mutation was subsequently found in all the 20 analyzed Finnish AGU patients, in the heterozygous form in all 53 carriers and in none of 67 control individuals, suggesting that it represents the major AGU causing mutation enriched in this isolated population. Since the mutation produces a change in the predicted flexibility of the AGA polypeptide chain and removes an intramolecular S-S bridge, it most probably explains the deficient enzyme activity found in cells and tissues of AGU patients.
Topics: Amino Acid Sequence; Animals; Aspartylglucosaminuria; Aspartylglucosylaminase; Base Sequence; Carbohydrate Metabolism, Inborn Errors; Cell Line; Chromosomes, Human, Pair 4; Cloning, Molecular; DNA; Female; Humans; Male; Molecular Sequence Data; Mutation; Oligonucleotide Probes; Pedigree; Polymerase Chain Reaction; Protein Conformation; RNA; Transfection
PubMed: 1703489
DOI: 10.1002/j.1460-2075.1991.tb07920.x -
Scientific Reports Nov 2016Aspartylglucosaminuria (AGU) is a lysosomal storage disorder that is caused by genetic deficiency of the enzyme aspartylglucosaminidase (AGA) which is involved in...
Aspartylglucosaminuria (AGU) is a lysosomal storage disorder that is caused by genetic deficiency of the enzyme aspartylglucosaminidase (AGA) which is involved in glycoprotein degradation. AGU is a progressive disorder that results in severe mental retardation in early adulthood. No curative therapy is currently available for AGU. We have here characterized the consequences of a novel AGU mutation that results in Thr122Lys exchange in AGA, and compared this mutant form to one carrying the worldwide most common AGU mutation, AGU-Fin. We show that T122K mutated AGA is expressed in normal amounts and localized in lysosomes, but exhibits low AGA activity due to impaired processing of the precursor molecule into subunits. Coexpression of T122K with wildtype AGA results in processing of the precursor into subunits, implicating that the mutation causes a local misfolding that prevents the precursor from becoming processed. Similar data were obtained for the AGU-Fin mutant polypeptide. We have here also identified small chemical compounds that function as chemical or pharmacological chaperones for the mutant AGA. Treatment of patient fibroblasts with these compounds results in increased AGA activity and processing, implicating that these substances may be suitable for chaperone mediated therapy for AGU.
Topics: Amino Acid Sequence; Aspartylglucosaminuria; Aspartylglucosylaminase; Base Sequence; Endoplasmic Reticulum; Fibroblasts; Golgi Apparatus; HEK293 Cells; HeLa Cells; Humans; Lysosomes; Male; Molecular Chaperones; Mutant Proteins; Mutation; Small Molecule Libraries
PubMed: 27876883
DOI: 10.1038/srep37583 -
JIMD Reports Sep 2021Aspartylglucosaminuria (AGU) (OMIM #208400) is a recessively inherited disorder of glycoprotein catabolism, a subset of the lysosomal storage disorders (LSDs)....
Aspartylglucosaminuria (AGU) (OMIM #208400) is a recessively inherited disorder of glycoprotein catabolism, a subset of the lysosomal storage disorders (LSDs). Deficiency of the enzyme (E.C. 3.5.1.26) leads to accumulation of aspartylglucosamine in various organs and its excretion in the urine. The disease is characterized by an initial period of normal development in infancy, a plateau in childhood, and subsequent regression in adolescence and adulthood. No curative treatments are currently available, leading to a protracted period of significant disability prior to early death. Hematopoietic stem cell transplantation (HSCT) has demonstrated efficacy in other LSDs, by providing enzyme replacement therapy in somatic viscera and decreasing substrate accumulation. Moreover, donor-derived monocytes cross the blood-brain barrier, differentiate into microglia, and secrete enzyme in the central nervous system (CNS). This has been shown to improve neurocognitive outcomes in other LSDs. The evidence to date for HSCT in AGU is varied, with marked improvement in glycosylasparaginase enzyme activity in the CNS in mice models, but varying neurocognitive outcomes in humans. We present a case series of four children with AGU who underwent HSCT at different ages (9 years, 5 years, 5 months, and 7 months of age), with long-term follow-up post-transplant (over 10 years). These cases demonstrate similar neurodevelopmental heterogeneity based on formal developmental assessments. The third case, transplanted prior to the onset of neurocognitive involvement, is developing normally despite a severe phenotype in other family members. This suggests that further research should examine the role of early HSCT in management of AGU.
PubMed: 34485011
DOI: 10.1002/jmd2.12222 -
The Biochemical Journal Mar 1976The isolation of liver N-aspartyl-beta-glucosaminidase in human aspartylglucosaminuria, where this enzyme activity is diminished, yields an enzyme molecule with the same...
The isolation of liver N-aspartyl-beta-glucosaminidase in human aspartylglucosaminuria, where this enzyme activity is diminished, yields an enzyme molecule with the same molecular weight and pH optimum as the normal enzyme. Its activity is 10% of that of the control preparation. Combination of both enzymes results in the summation of both activities, and the pathological enzyme does not inhibit the control preparation. It is concluded that no change into a totally different isoenzyme has occurred in aspartylglucosaminuria.
Topics: Amidohydrolases; Aspartylglucosaminuria; Aspartylglucosylaminase; Electrophoresis, Polyacrylamide Gel; Humans; Liver
PubMed: 942387
DOI: 10.1042/bj1530749 -
Orphanet Journal of Rare Diseases Feb 2014Oligosaccharidoses, which belong to the lysosomal storage diseases, are inherited metabolic disorders due to the absence or the loss of function of one of the enzymes...
BACKGROUND
Oligosaccharidoses, which belong to the lysosomal storage diseases, are inherited metabolic disorders due to the absence or the loss of function of one of the enzymes involved in the catabolic pathway of glycoproteins and indirectly of glycosphingolipids. This enzymatic deficiency typically results in the abnormal accumulation of uncompletely degraded oligosaccharides in the urine. Since the clinical features of many of these disorders are not specific for a single enzyme deficiency, unambiguous screening is critical to limit the number of costly enzyme assays which otherwise must be performed.
METHODS
Here we provide evidence for the advantages of using a MALDI-TOF/TOF (matrix-assisted laser desorption ionization time-of-flight) mass spectrometric (MS) method for screening oligosaccharidoses. Urine samples from previously diagnosed patients or from unaffected subjects were randomly divided into a training set and a blind testing set. Samples were directly analyzed without prior treatment.
RESULTS
The characteristic MS and MS/MS molecular profiles obtained allowed us to identify fucosidosis, aspartylglucosaminuria, GM1 gangliosidosis, Sandhoff disease, α-mannosidosis, sialidosis and mucolipidoses type II and III.
CONCLUSIONS
This method, which is easily run in less than 30 minutes, is performed in a single step, and is sensitive and specific. Invaluable for clinical chemistry purposes this MALDI-TOF/TOF mass spectrometry procedure is semi-automatizable and suitable for the urinary screening of oligosacharidoses.
Topics: Female; Humans; Lysosomal Storage Diseases; Male; Oligosaccharides; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tandem Mass Spectrometry
PubMed: 24502792
DOI: 10.1186/1750-1172-9-19 -
Journal of Medical Genetics May 1999Aspartylglucosaminuria (AGU) is a lysosomal storage disorder caused by deficiency of aspartylglucosaminidase (AGA). The main symptom is progressive mental retardation. A...
Aspartylglucosaminuria (AGU) is a lysosomal storage disorder caused by deficiency of aspartylglucosaminidase (AGA). The main symptom is progressive mental retardation. A spectrum of different mutations has been reported in this disease, one missense mutation (Cys163Ser) being responsible for the majority of Finnish cases. We were able to examine 66 Finnish AGU patients for changes in the oral mucosa and 44 of these for changes in facial skin. Biopsy specimens of 16 oral lesions, 12 of them associated with the teeth, plus two facial lesions were studied histologically. Immunohistochemical staining for AGA was performed on 15 oral specimens. Skin was seborrhoeic in adolescent and adult patients, with erythema of the facial skin already common in childhood. Of 44 patients, nine (20%) had facial angiofibromas, tumours primarily occurring in association with tuberous sclerosis. Oedemic buccal mucosa (leucoedema) and gingival overgrowths were more frequent in AGU patients than in controls (p<0.001). Of 16 oral mucosal lesions studied histologically, 15 represented fibroepithelial or epithelial hyperplasias and were reactive in nature. Cytoplasmic vacuolisation was evident in four. Immunohistochemically, expression of AGA in AGU patients' mucosal lesions did not differ from that seen in corresponding lesions of normal subjects. Thus, the high frequency of mucosal overgrowth in AGU patients does not appear to be directly associated with lysosomal storage or with alterations in the level of AGA expression.
Topics: Acetylglucosamine; Adolescent; Adult; Angiofibroma; Aspartylglucosaminuria; Aspartylglucosylaminase; Child; Child, Preschool; Face; Facial Neoplasms; Fibroma; Finland; Gingiva; Humans; Immunohistochemistry; Lysosomal Storage Diseases; Middle Aged; Mouth Mucosa; Skin; Skin Neoplasms
PubMed: 10353787
DOI: No ID Found