A contradictory treatment for lysosomal storage disorders: inhibitors enhance mutant enzyme activity

Fan, Jintu

doi:10.1016/s0165-6147(03)00158-5

Cited by 218 publications

(130 citation statements)

References 55 publications

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“…For example, unstable variants of lysosomal ␣-galactosidase A, associated with Fabry disease, were rescued by addition of a competitive inhibitor, 1-deoxygalactonorijimycin (42,43). In other studies, increased protein levels were seen following treatment of cells expressing mutant acid ␤-glucosidase (defective in Gaucher disease) or a V2 vasopressin receptor mutant (associated with nephrogenic diabetes insipidus) with active site inhibitors (44,45). These observations, along with our results with AGT-PLP-AOA, suggest that an AGT-specific inhibitor could be therapeutically useful for PH1, particularly when used in conjunction with PLP.…”

Section: Discussionmentioning

confidence: 99%

In Vivo and in Vitro Examination of Stability of Primary Hyperoxaluria-associated Human Alanine:Glyoxylate Aminotransferase

Hopper

Pittman

Fitzgerald

et al. 2008

Journal of Biological Chemistry

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Primary hyperoxaluria type I is a severe kidney stone disease caused by mutations in the protein alanine:glyoxylate aminotransferase (AGT). Many patients have mutations in AGT that are not deleterious alone but act synergistically with a common minor allele polymorphic variant to impair protein folding, dimerization, or localization. Although studies suggest that the minor allele variant itself is destabilized, no direct stability studies have been carried out. In this report, we analyze AGT function and stability using three approaches. First, we describe a yeast complementation growth assay for AGT, in which we show that human AGT can substitute for function of yeast Agx1 and that mutations associated with disease in humans show reduced growth in yeast. The reduced growth of minor allele mutants reflects reduced protein levels, indicating that these proteins are less stable than wild-type AGT in yeast. We further examine stability of AGT alleles in vitro using two direct methods, a mass spectrometry-based technique (stability of unpurified proteins from rates of H/D exchange) and differential scanning fluorimetry. We also examine the effect of known ligands pyridoxal 5-phosphate and aminooxyacetic acid on stability. Our work establishes that the minor allele is destabilized and that pyridoxal 5-phosphate and aminooxyacetic acid binding significantly stabilizes both alleles. To our knowledge, this is the first work that directly measures relative stabilities of AGT variants and ligand complexes. Because previous studies suggest that stabilizing compounds (i.e. pharmacological chaperones) may be effective for treatment of primary hyperoxaluria, we propose that the methods described here can be used in high throughput screens for compounds that stabilize AGT mutants.Deficiencies in the enzyme alanine:glyoxylate aminotransferase (AGT) 3 cause primary hyperoxaluria type I (PH1), a severe autosomal recessive kidney stone disease (1, 2). In humans, AGT is responsible for conversion of glyoxylate to glycine in the liver. Without functional AGT, glyoxylate builds up and is converted to calcium oxalate, which is deposited in the kidneys and can lead to kidney stones and renal failure. In many patients, deficiency of AGT results from one or two amino acid changes that decrease the stability of this enzyme. As a result, AGT may be degraded, become improperly localized, or form nonfunctional aggregates (1, 2).Over 50 different mutations of AGT and two polymorphic variants have been identified (1, 3). The two allelic forms consist of a "wild-type" major allele, AGTma, and a minor allele, AGTmi. The minor allele is present in ϳ20% of European and North American populations and contains P11L and I340M substitutions in the amino acid sequence and a 74-bp duplication in intron 1 (4, 5). The P11L and I340M substitutions, particularly P11L, have several biochemical effects, including decreasing catalytic activity and slowing the dimerization rate, but by themselves are not disease-causing (4, 5). The minor allele of AGT is delete...

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Section: Discussionmentioning

confidence: 99%

In Vivo and in Vitro Examination of Stability of Primary Hyperoxaluria-associated Human Alanine:Glyoxylate Aminotransferase

Hopper

Pittman

Fitzgerald

et al. 2008

Journal of Biological Chemistry

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“…This also serves as proof of concept that the GlcCerase structure can be used as a starting point for designing structure-based drugs aimed at restoring the activity of defective GlcCerase (3). Important among these are the small molecular weight chaperones that, upon binding to the active site, restore trafficking of improperly folded GlcCerase out of the endoplasmic reticulum and through the secretory pathway to lysosomes, thereby restoring normal, or near-normal, lysosomal GlcCerase levels (5,26).…”

Section: Discussionmentioning

confidence: 99%

X-ray Structure of Human Acid-β-Glucosidase Covalently Bound to Conduritol-B-Epoxide

Premkumar

Sawkar

Boldin-Adamsky

et al. 2005

Journal of Biological Chemistry

102

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Gaucher disease is an inherited metabolic disorder caused by mutations in the lysosomal enzyme acid-␤-glucosidase (GlcCerase). We recently determined the x-ray structure of GlcCerase to 2.0 Å resolution ( -349 and 394 -399) located at the entrance to the active site in native GlcCerase is observed in the GlcCerase-CBE structure, a conformation in which the active site is accessible to CBE. Analysis of the dynamics of these two alternative conformations suggests that the two loops act as a lid at the entrance to the active site. This possibility is supported by a cluster of mutations in loop 394 -399 that cause Gaucher disease by reducing catalytic activity. Moreover, in silico mutational analysis demonstrates that all these mutations stabilize the conformation that limits access to the active site, thus providing a mechanistic explanation of how mutations in this loop result in Gaucher disease.

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“…If the expression studies confirm this hypothesis, active site-specific inhibitors of the enzyme could be tested for their ability to stabilize the mutants and modify their conformation closer to that of the wild-type enzyme, increasing the level of their residual activity as has been demonstrated for other lysosomal enzymes (reviewed in Fan [2003]; Desnick [2004]). Together with the inhibitors of heparan sulfate synthesis [Piotrowska et al, 2006;Jakó bkiewicz-Banecka et al, 2007], such ''pharmacological chaperones'' may be an interesting subject of future research in this area.…”

Section: Clinical and Diagnostic Relevancementioning

confidence: 93%

Sanfilippo syndrome type C: mutation spectrum in the heparan sulfate acetyl-CoA: α-glucosaminide N-acetyltransferase (HGSNAT) gene

et al. 2009

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Mucopolysaccharidosis (MPS) type IIIC or Sanfilippo syndrome type C is a rare autosomal recessive disorder caused by the deficiency of the lysosomal membrane enzyme, heparan sulfate acetyl-CoA (AcCoA): a-glucosaminide N-acetyltransferase (HGSNAT; EC 2.3.1.78), which catalyzes transmembrane acetylation of the terminal glucosamine residues of heparan sulfate prior to their hydrolysis by a-N-acetylglucosaminidase. Lysosomal storage of undegraded heparan sulfate in the cells of affected patients leads to neuronal death, causing neurodegeneration and severely impaired development accompanied by mild visceral and skeletal abnormalities, including mild dwarfism, coarse facies, and joint stiffness. To date, 50 HGSNAT mutations have been identified in MPS IIIC patients: 40 were previously published and 10 novel mutations are reported here. The mutations span the entire structure of the gene and include 13 splice-site mutations, 11 insertions and deletions, 8 nonsense mutations, and 18 missense mutations (http://chromium.liacs.nl/LOVD2/home.php? select_db 5 HGSNAT). In addition, four polymorphisms result in amino acid changes that do not affect activity of the enzyme. In this work we discuss the spectrum of MPS IIIC mutations, their clinical presentation and distribution within the patient population, and speculate how the mutations may affect the structure and function of HGSNAT. Hum Mutat 30,[918][919][920][921][922][923][924][925]

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A contradictory treatment for lysosomal storage disorders: inhibitors enhance mutant enzyme activity

Cited by 218 publications

References 55 publications

In Vivo and in Vitro Examination of Stability of Primary Hyperoxaluria-associated Human Alanine:Glyoxylate Aminotransferase

In Vivo and in Vitro Examination of Stability of Primary Hyperoxaluria-associated Human Alanine:Glyoxylate Aminotransferase

X-ray Structure of Human Acid-β-Glucosidase Covalently Bound to Conduritol-B-Epoxide

Sanfilippo syndrome type C: mutation spectrum in the heparan sulfate acetyl-CoA: α-glucosaminide N-acetyltransferase (HGSNAT) gene

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