2byl
From Proteopedia
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|SITE= <scene name='pdbsite=AC1:Plp+(Pyridoxal-5'-Phosphate)+Is+Bound+To+The+Protein+Via+...'>AC1</scene> | |SITE= <scene name='pdbsite=AC1:Plp+(Pyridoxal-5'-Phosphate)+Is+Bound+To+The+Protein+Via+...'>AC1</scene> | ||
|LIGAND= <scene name='pdbligand=PLP:PYRIDOXAL-5'-PHOSPHATE'>PLP</scene> | |LIGAND= <scene name='pdbligand=PLP:PYRIDOXAL-5'-PHOSPHATE'>PLP</scene> | ||
| - | |ACTIVITY= [http://en.wikipedia.org/wiki/Ornithine_aminotransferase Ornithine aminotransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.6.1.13 2.6.1.13] | + | |ACTIVITY= <span class='plainlinks'>[http://en.wikipedia.org/wiki/Ornithine_aminotransferase Ornithine aminotransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.6.1.13 2.6.1.13] </span> |
|GENE= | |GENE= | ||
| + | |DOMAIN= | ||
| + | |RELATEDENTRY= | ||
| + | |RESOURCES=<span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=2byl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=2byl OCA], [http://www.ebi.ac.uk/pdbsum/2byl PDBsum], [http://www.rcsb.org/pdb/explore.do?structureId=2byl RCSB]</span> | ||
}} | }} | ||
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==Overview== | ==Overview== | ||
Ornithine aminotransferase and 4-aminobutyrate aminotransferase are related pyridoxal phosphate-dependent enzymes having different substrate specificities. The atomic structures of these enzymes have shown (i) that active site differences are limited to the steric positions occupied by two tyrosine residues in ornithine aminotransferase and (ii) that, uniquely among related, structurally characterized aminotransferases, the conserved arginine that binds the alpha-carboxylate of alpha-amino acids interacts tightly with a glutamate residue. To determine the contribution of these residues to the specificities of the enzymes, we analyzed site-directed mutants of ornithine aminotransferase by rapid reaction kinetics, x-ray crystallography, and 13C NMR spectroscopy. Mutation of one tyrosine (Tyr-85) to isoleucine, as found in aminobutyrate aminotransferase, decreased the rate of the reaction of the enzyme with ornithine 1000-fold and increased that with 4-aminobutyrate 16-fold, indicating that Tyr-85 is a major determinant of specificity toward ornithine. Unexpectedly, the limiting rate of the second half of the reaction, conversion of ketoglutarate to glutamate, was greatly increased, although the kinetics of the reverse reaction were unaffected. A mutant in which the glutamate (Glu-235) that interacts with the conserved arginine was replaced by alanine retained its regiospecificity for the delta-amino group of ornithine, but the glutamate reaction was enhanced 650-fold, whereas only a 5-fold enhancement of the ketoglutarate reaction rate resulted. A model is proposed in which conversion of the enzyme to its pyridoxamine phosphate form disrupts the internal glutamate-arginine interaction, thus enabling ketoglutarate but not glutamate to be a good substrate. | Ornithine aminotransferase and 4-aminobutyrate aminotransferase are related pyridoxal phosphate-dependent enzymes having different substrate specificities. The atomic structures of these enzymes have shown (i) that active site differences are limited to the steric positions occupied by two tyrosine residues in ornithine aminotransferase and (ii) that, uniquely among related, structurally characterized aminotransferases, the conserved arginine that binds the alpha-carboxylate of alpha-amino acids interacts tightly with a glutamate residue. To determine the contribution of these residues to the specificities of the enzymes, we analyzed site-directed mutants of ornithine aminotransferase by rapid reaction kinetics, x-ray crystallography, and 13C NMR spectroscopy. Mutation of one tyrosine (Tyr-85) to isoleucine, as found in aminobutyrate aminotransferase, decreased the rate of the reaction of the enzyme with ornithine 1000-fold and increased that with 4-aminobutyrate 16-fold, indicating that Tyr-85 is a major determinant of specificity toward ornithine. Unexpectedly, the limiting rate of the second half of the reaction, conversion of ketoglutarate to glutamate, was greatly increased, although the kinetics of the reverse reaction were unaffected. A mutant in which the glutamate (Glu-235) that interacts with the conserved arginine was replaced by alanine retained its regiospecificity for the delta-amino group of ornithine, but the glutamate reaction was enhanced 650-fold, whereas only a 5-fold enhancement of the ketoglutarate reaction rate resulted. A model is proposed in which conversion of the enzyme to its pyridoxamine phosphate form disrupts the internal glutamate-arginine interaction, thus enabling ketoglutarate but not glutamate to be a good substrate. | ||
| - | |||
| - | ==Disease== | ||
| - | Known disease associated with this structure: Gyrate atrophy of choroid and retina with ornithinemia, B6 responsive or unresponsive OMIM:[[http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=258870 258870]] | ||
==About this Structure== | ==About this Structure== | ||
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[[Category: Peneff, C.]] | [[Category: Peneff, C.]] | ||
[[Category: Schirmer, T.]] | [[Category: Schirmer, T.]] | ||
| - | [[Category: PLP]] | ||
[[Category: aminotransferase]] | [[Category: aminotransferase]] | ||
[[Category: disease mutation]] | [[Category: disease mutation]] | ||
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[[Category: transit peptide]] | [[Category: transit peptide]] | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Mar 31 02:13:46 2008'' |
Revision as of 23:13, 30 March 2008
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| , resolution 2.15Å | |||||||
|---|---|---|---|---|---|---|---|
| Sites: | |||||||
| Ligands: | |||||||
| Activity: | Ornithine aminotransferase, with EC number 2.6.1.13 | ||||||
| Resources: | FirstGlance, OCA, PDBsum, RCSB | ||||||
| Coordinates: | save as pdb, mmCIF, xml | ||||||
STRUCTURE OF ORNITHINE AMINOTRANSFERASE TRIPLE MUTANT Y85I Y55A G320F
Overview
Ornithine aminotransferase and 4-aminobutyrate aminotransferase are related pyridoxal phosphate-dependent enzymes having different substrate specificities. The atomic structures of these enzymes have shown (i) that active site differences are limited to the steric positions occupied by two tyrosine residues in ornithine aminotransferase and (ii) that, uniquely among related, structurally characterized aminotransferases, the conserved arginine that binds the alpha-carboxylate of alpha-amino acids interacts tightly with a glutamate residue. To determine the contribution of these residues to the specificities of the enzymes, we analyzed site-directed mutants of ornithine aminotransferase by rapid reaction kinetics, x-ray crystallography, and 13C NMR spectroscopy. Mutation of one tyrosine (Tyr-85) to isoleucine, as found in aminobutyrate aminotransferase, decreased the rate of the reaction of the enzyme with ornithine 1000-fold and increased that with 4-aminobutyrate 16-fold, indicating that Tyr-85 is a major determinant of specificity toward ornithine. Unexpectedly, the limiting rate of the second half of the reaction, conversion of ketoglutarate to glutamate, was greatly increased, although the kinetics of the reverse reaction were unaffected. A mutant in which the glutamate (Glu-235) that interacts with the conserved arginine was replaced by alanine retained its regiospecificity for the delta-amino group of ornithine, but the glutamate reaction was enhanced 650-fold, whereas only a 5-fold enhancement of the ketoglutarate reaction rate resulted. A model is proposed in which conversion of the enzyme to its pyridoxamine phosphate form disrupts the internal glutamate-arginine interaction, thus enabling ketoglutarate but not glutamate to be a good substrate.
About this Structure
2BYL is a Single protein structure of sequence from Homo sapiens. Full crystallographic information is available from OCA.
Reference
Determinants of substrate specificity in omega-aminotransferases., Markova M, Peneff C, Hewlins MJ, Schirmer T, John RA, J Biol Chem. 2005 Oct 28;280(43):36409-16. Epub 2005 Aug 11. PMID:16096275
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