Alanine : glyoxylate aminotransferase of <i>Saccharomyces cerevisiae</i>–encoding gene <i>AGX1</i> and metabolic significance

Schlösser, Thomas; Gätgens, Cornelia; Weber, U.; Stahmann, K.‐Peter

doi:10.1002/yea.1058

Cited by 32 publications

(26 citation statements)

References 43 publications

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“…Although AGX1 is not essential for viability, yeast lacking AGX1 in a shm1, shm2, and gly1 background are unable to grow on medium that contains ethanol unless exogenous glycine is provided (16). Plasmid-expressed yeast Agx1 protein can restore ethanol-dependent growth to an agx1-deficient strain, YCG-F r (16). The Agx1 enzyme has 23% identity to human AGT, with residues that are conserved among all AGTs, such as a common pyridoxal 5Ј-phosphate-binding site (Fig.…”

Section: Expression Of Agt Disease Variants In Yeast-yeast Alaninementioning

confidence: 99%

“…We examined whether expression of plasmid-derived human AGT could complement growth of yeast lacking AGX1 in the YCG-F r strain (16). This strain is unable to grow on glycine-free medium, because it cannot synthesize glycine endogenously (16).…”

Section: Expression Of Agt Disease Variants In Yeast-yeast Alaninementioning

confidence: 99%

“…This strain is unable to grow on glycine-free medium, because it cannot synthesize glycine endogenously (16). When streaked on plates containing ethanol, YCG-F r cells expressing Agx1 (p416ADH-Agx1) are able to grow, because the Agx1 enzyme converts glyoxylate to glycine (Fig.…”

Section: Expression Of Agt Disease Variants In Yeast-yeast Alaninementioning

confidence: 99%

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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: Expression Of Agt Disease Variants In Yeast-yeast Alaninementioning

confidence: 99%

Section: Expression Of Agt Disease Variants In Yeast-yeast Alaninementioning

confidence: 99%

See 1 more Smart Citation

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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“…In yeast, disruption of AGT can lead to glycine auxo-trophy. For example, a yeast AGT deletion strain was unable to grow when glucose was the only carbon source, unless glycine was supplemented (Schlosser et al, 2004). The importance of hepatic AGT in minimizing endogenous oxalate production in some mammals is clearly shown by the autosomal recessive disorder of glyoxylate metabolism known as primary hyperoxaluria type 1.…”

Section: Primary Function Of Agtmentioning

confidence: 99%

“…aegypti AGT showed that this AGT, unlike mammalian AGTs, functions virtually exclusively on the glyoxylate to glycine pathway (i.e., functions as a true AGT) (Han et al, 2006). It has been shown that AGT is essential in human and yeast (Danpure et al, 1996;Danpure, 2001;Schlosser et al, 2004) and it likely is crucial in insects. In our previous study, we demonstrated that Ae.…”

Section: Possible Driving Force For Mosquito Agt Functional Adaptatiomentioning

confidence: 99%

The tryptophan oxidation pathway in mosquitoes with emphasis on xanthurenic acid biosynthesis

Han¹,

Beerntsen²,

Li³

2007

Journal of Insect Physiology

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Oxidation of tryptophan to kynurenine and 3-hydroxykynurenine (3-HK) is the major catabolic pathway in mosquitoes. However, 3-HK is oxidized easily under physiological conditions, resulting in the production of reactive radical species. To overcome this problem, mosquitoes have developed an efficient mechanism to prevent 3-HK from accumulating by converting this chemically reactive compound to the chemically stable xanthurenic acid. Interestingly, 3-HK is a precursor for the production of compound eye pigments during the pupal and early adult stages; consequently, mosquitoes need to preserve and transport 3-HK for compound eye pigmentation in pupae and adults. This review summarizes the tryptophan oxidation pathway, compares and contrasts the mosquito tryptophan oxidation pathway with other model species, and discusses possible driving forces leading to the functional adaptation and evolution of enzymes involved in the mosquito tryptophan oxidation pathway.

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Amino Acid Biosynthesis

Parker

Pratt

2010

Amino Acids, Peptides and Proteins in Organic Chemistry

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The ribosomal synthesis of proteins utilizes a family of 20 a-amino acids that are universally coded by the translation machinery; in addition, two further a-amino acids, selenocysteine and pyrrolysine, are now believed to be incorporated into proteins via ribosomal synthesis in some organisms. More than 300 other amino acid residues have been identified in proteins, but most are of restricted distribution and produced via post-translational modification of the ubiquitous protein amino acids [1]. The ribosomally encoded a-amino acids described here ultimately derive from a-keto acids by a process corresponding to reductive amination. The most important biosynthetic distinction relates to whether appropriate carbon skeletons are pre-existing in basic metabolism or whether they have to be synthesized de novo and this division underpins the structure of this chapter.There are a small number of a-keto acids ubiquitously found in core metabolism, notably pyruvate (and a related 3-phosphoglycerate derivative from glycolysis), together with two components of the tricarboxylic acid cycle (TCA), oxaloacetate and a-ketoglutarate (a-KG). These building blocks ultimately provide the carbon skeletons for unbranched a-amino acids of three, four, and five carbons, respectively. a-Amino acids with shorter (glycine) or longer (lysine and pyrrolysine) straight chains are made by alternative pathways depending on the available raw materials. The strategic challenge for the biosynthesis of most straight-chain amino acids centers around two issues: how is the a-amino function introduced into the carbon skeleton and what functional group manipulations are required to generate the diversity of side-chain functionality required for the protein function?The core family of straight-chain amino acids does not provide all the functionality required for proteins. a-Amino acids with branched side-chains are used for two purposes; the primary need is related to protein structural issues. Proteins fold into well-defined three-dimensional shapes by virtue of their amphipathic nature: a significant fraction of the amino acid side-chains are of low polarity and the hydrophobic effect drives the formation of ordered structures in which these side-chains are buried away from water. In contrast to the straight-chain amino

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Alanine : glyoxylate aminotransferase of Saccharomyces cerevisiae–encoding gene AGX1 and metabolic significance

Cited by 32 publications

References 43 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

The tryptophan oxidation pathway in mosquitoes with emphasis on xanthurenic acid biosynthesis

Amino Acid Biosynthesis

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