Identification of cysteine-319 as the target amino acid of 8-[(4-bromo-2,3-dioxobutyl)thio]adenosine 5'-triphosphate in bovine liver glutamate dehydrogenase

Ozturk, Derya H.; Colman, Roberta F.

doi:10.1021/bi00243a013

Cited by 13 publications

(17 citation statements)

References 44 publications

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“…Similar results with discrepancy using classical chemical probes were also reported by the same research group to identify other regulatory sites within bovine liver GDH. The NADH binding site was proposed to be modified by an ATP analogue at Cys 319 (9), by a GMP probe at Met 169 and Tyr 262 (10), and by the adenosine analogue at Lys 420 and Tyr 190 (11). The GTP binding site was also proposed to be modified by 5Ј-p-(fluorosulfonyl)benzoyl-1,N 6 -ethenoadenosine at Tyr 262 (24).…”

Section: Discussionmentioning

confidence: 99%

“…This is especially likely if they display low affinity for the binding site being studies. Their lack of specificity may be the reason for the wide three-dimensional distribution of the residues identified using classical chemical probes as being in the NADH inhibitory site of GDH (9,10). An analysis of the three-dimensional structure of the mammalian enzyme should supplement the understanding of the nature and location of these regulatory sites.…”

Section: Discussionmentioning

confidence: 99%

“…The studies using classical chemical probes to identify the NADH and GTP binding site within bovine liver GDH, however, gave a wide scatter of modified residues throughout most of the proposed three-dimensional structure of GDH. For instance, the NADH binding site was proposed to be modified by an ATP analogue at Cys 319 (9), by a GMP probe at Met 169 and Tyr 262 (10), and by the adenosine analogue at Lys 420 and Tyr 190 (11). It seems, therefore, that the base moiety has not been effective at directing the site of modification by classical chemical probes.…”

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confidence: 99%

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Photoaffinity Labeling of Brain Glutamate Dehydrogenase Isoproteins with an Azido-ADP

Cho

Yoon

1999

Journal of Biological Chemistry

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The ADP binding site within two types of bovine brain glutamate dehydrogenase isoproteins (GDH I and GDH II) was identified using photoaffinity labeling with [ Glutamate dehydrogenase (GDH 1 ; EC 1.4.1.3) is a family of enzymes that catalyze the reversible deamination of L-glutamate to 2-oxoglutarate using NAD ϩ , NADP ϩ , or both as coenzyme (1). Since the pathology of the disorders associated with GDH defects is restricted to the brain, the enzyme may be of particular importance in the biology of the nervous system. The importance of the pathophysiological nature of the GDH-deficient neurological disorders has attracted considerable interest (2). The enzyme isolated from one of the patients with a variant form of multisystem atrophy displayed a marked reduction of one of the GDH isoproteins (3). Although the origin of the GDH polymorphism is not known, it has been reported that the presence of differently sized mRNAs and multiple gene copies for GDH occur in the human brain (4). A novel cDNA encoded by an X chromosome-linked intronless gene also has been isolated from human retina (5). However, it is not known whether the distinct properties of the GDH isoproteins are essential for the regulation of glutamate metabolism. Therefore, it is essential to have a detailed structural and functional description of the various types of brain GDH to elucidate the pathophysiological nature of the GDH-deficient neurological disorders.Mammalian GDH is composed of six identical subunits, and the regulation of GDH is very complex (1). It has been a major goal to identify the substrate and regulatory binding sites of GDH. It is the only recent years that the three-dimensional structure of GDH from microorganisms is available (6, 7). Recently, crystallization of bovine liver GDH was reported for the first time from the mammalian sources (8). However, remarkably little is known about the detailed structure of mammalian GDH, especially the brain enzymes. Although regulatory and substrate binding sites of GDH have been reported, the results are quite controversial. Several classical chemical probes have been used to attempt resolution of these binding sites. The studies using classical chemical probes to identify the NADH and GTP binding site within bovine liver GDH, however, gave a wide scatter of modified residues throughout most of the proposed three-dimensional structure of GDH. For instance, the NADH binding site was proposed to be modified by an ATP analogue at Cys 319 (9), by a GMP probe at Met 169 and Tyr 262(10), and by the adenosine analogue at Lys 420 and Tyr 190 (11). It seems, therefore, that the base moiety has not been effective at directing the site of modification by classical chemical probes.Alternatively, identifying nucleotide binding sites of variety of proteins has been advanced by the use of nucleotide photoaffinity analogues that selectively insert into a site upon photoactivation with ultraviolet light. Stanley et al. (23) have reported that the hyperinsulinism-hyperammonemia syndrome is caused by mutat...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Photoaffinity Labeling of Brain Glutamate Dehydrogenase Isoproteins with an Azido-ADP

Cho

Yoon

1999

Journal of Biological Chemistry

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“…This is especially likely if they display low affinity for the binding site being studied. Their lack of specificity may be the reason for the wide three-dimensional distribution of the residues identified using classical chemical probes as being in the NADH inhibitory site of GDH (8,9). Very recently, Stanley et al (38) have reported that the hyperinsulinism-hyperammonemia syndrome is caused by mutations in GDH gene that affect enzyme sensitivity to GTPinduced inhibition.…”

Section: Nad ϩ Binding Site Of Brain Gdh Isoproteinsmentioning

confidence: 99%

“…The studies using classical chemical probes to identify the NADH and GTP binding sites within bovine liver GDH, however, gave a wide scatter of modified residues throughout most of the proposed threedimensional structure of GDH. For instance, the NADH binding site was proposed to be modified by an ATP analogue at Cys 319 (8), by a GMP probe at Met 169 and Tyr 262 (9), and by the adenosine analogue at Lys 420 and Tyr 190 (10). It seems, therefore, that the base moiety has not been effective at directing the site of modification by classical chemical probes.…”

mentioning

confidence: 99%

Identification of an NAD+ Binding Site of Brain Glutamate Dehydrogenase Isoproteins by Photoaffinity Labeling

Cho

Yoon

Ahn

et al. 1998

Journal of Biological Chemistry

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Photoaffinity labeling with [32 P]nicotinamide 2-azidoadenosine dinucleotide (2N 3 NAD ؉ ) was used to identify the NAD ؉ binding site within two types of glutamate dehydrogenase isoproteins (GDH I and GDH II) isolated from bovine brain. In the absence of photolysis, 2N 3 NAD ؉ is a substrate for the GDH isoproteins. When the enzymes were covalently modified by photolysis in the presence of saturating amounts of photoprobe, about 50% inhibition of the GDH activities was observed. Photoinsertion of probe was increased by GTP or glutarate and decreased by NAD ؉ or ADP. With the combination of immobilized boronate affinity chromatography and reversed-phase HPLC, photolabelcontaining peptides generated with trypsin were isolated. This identified a portion of the adenine ring binding domain of GDH isoproteins as the region containing the sequence, CIAVGXSDGSIWNPDGIDPK for both GDH isoproteins, corresponding to Cys 270 through Lys 289 of the amino acid sequence of well known bovine liver GDH. The X indicates a position for which no phenylthiohydantoin-derivative could be assigned. The missing residue, however, can be designated as a photolabeled glutamate since the sequences including the glutamate residue in question have a complete identity with those of the other GDH species known. Photolabeling of these peptides was prevented by the presence of NAD ؉ during photolysis. These results demonstrate selectivity of the photoprobe for the NAD ؉ binding site and suggest that the peptide identified using the photoprobe is located in the NAD ؉ binding domain of the brain GDH isoproteins. Both amino acid sequencing and compositional analysis identified Glu 275 as the site of photoinsertion.

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Identification of Tyr115 labeled by S-(4-bromo-2,3-dioxobutyl)glutathione in the hydrophobic substrate binding site of glutathione S-transferase, isoenzyme 3-3

Katusz

Bono

Colman

1992

Archives of Biochemistry and Biophysics

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Identification of cysteine-319 as the target amino acid of 8-[(4-bromo-2,3-dioxobutyl)thio]adenosine 5'-triphosphate in bovine liver glutamate dehydrogenase

Cited by 13 publications

References 44 publications

Photoaffinity Labeling of Brain Glutamate Dehydrogenase Isoproteins with an Azido-ADP

Photoaffinity Labeling of Brain Glutamate Dehydrogenase Isoproteins with an Azido-ADP

Identification of an NAD+ Binding Site of Brain Glutamate Dehydrogenase Isoproteins by Photoaffinity Labeling

Identification of Tyr115 labeled by S-(4-bromo-2,3-dioxobutyl)glutathione in the hydrophobic substrate binding site of glutathione S-transferase, isoenzyme 3-3

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