1r8g
From Proteopedia
(New page: 200px<br /><applet load="1r8g" size="450" color="white" frame="true" align="right" spinBox="true" caption="1r8g, resolution 2.15Å" /> '''Structure and functi...) |
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| - | [[Image:1r8g.jpg|left|200px]]<br /><applet load="1r8g" size=" | + | [[Image:1r8g.jpg|left|200px]]<br /><applet load="1r8g" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1r8g, resolution 2.15Å" /> | caption="1r8g, resolution 2.15Å" /> | ||
'''Structure and function of YbdK'''<br /> | '''Structure and function of YbdK'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The Escherichia coli open reading frame YbdK encodes a member of a large | + | The Escherichia coli open reading frame YbdK encodes a member of a large bacterial protein family of unknown biological function. The sequences within this family are remotely related to the sequence of gamma-glutamate-cysteine ligase (gamma-GCS), an enzyme in the glutathione biosynthetic pathway. A gene encoding gamma-GCS in E. coli is already known. The 2.15 A resolution crystal structure of YbdK reveals an overall fold similar to that of glutamine synthetase (GS), a nitrogen metabolism enzyme that ligates glutamate and ammonia to yield glutamine. GS and gamma-GCS perform related chemical reactions and require ATP and Mg2+ for their activity. The Mg2+-dependent binding of ATP to YbdK was confirmed by fluorescence spectroscopy employing 2'(or 3')-O-(trinitrophenyl)adenosine 5'-triphosphate, and yielding a dissociation constant of 3 +/- 0.5 microM. The structure of YbdK contains a crevice that corresponds to the binding sites of ATP, Mg2+ and glutamate in GS. Many of the GS residues that coordinate the metal ions and interact with glutamic acid and the phosphoryl and ribosyl groups of ATP are also present in YbdK. GS amino acids that have been associated with ammonia binding have no obvious counterparts in YbdK, consistent with a substrate specificity that is different from that of GS. Ligase activity between glutamic acid and each of the twenty amino acid residues was tested on high performance liquid chromatography (HPLC) by following the hydrolysis of ATP to ADP. Catalysis was observed only with cysteine. A pyruvate kinase/lactic acid dehydrogenase coupled assay was used to rule out GS activity and to determine that YbdK exhibits gamma-GCS activity. The catalytic rate was found to be approximately 500-fold slower than that reported for authentic gamma-GCS. |
==About this Structure== | ==About this Structure== | ||
| - | 1R8G is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli]. Full crystallographic information is available from [http:// | + | 1R8G is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1R8G OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Lehmann, C.]] | [[Category: Lehmann, C.]] | ||
[[Category: Pullalarevu, S.]] | [[Category: Pullalarevu, S.]] | ||
| - | [[Category: S2F, Structure | + | [[Category: S2F, Structure 2.Function Project.]] |
[[Category: carboxylate-amine ligase]] | [[Category: carboxylate-amine ligase]] | ||
[[Category: hypothetical protein]] | [[Category: hypothetical protein]] | ||
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[[Category: ybdk]] | [[Category: ybdk]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 14:48:15 2008'' |
Revision as of 12:48, 21 February 2008
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Structure and function of YbdK
Overview
The Escherichia coli open reading frame YbdK encodes a member of a large bacterial protein family of unknown biological function. The sequences within this family are remotely related to the sequence of gamma-glutamate-cysteine ligase (gamma-GCS), an enzyme in the glutathione biosynthetic pathway. A gene encoding gamma-GCS in E. coli is already known. The 2.15 A resolution crystal structure of YbdK reveals an overall fold similar to that of glutamine synthetase (GS), a nitrogen metabolism enzyme that ligates glutamate and ammonia to yield glutamine. GS and gamma-GCS perform related chemical reactions and require ATP and Mg2+ for their activity. The Mg2+-dependent binding of ATP to YbdK was confirmed by fluorescence spectroscopy employing 2'(or 3')-O-(trinitrophenyl)adenosine 5'-triphosphate, and yielding a dissociation constant of 3 +/- 0.5 microM. The structure of YbdK contains a crevice that corresponds to the binding sites of ATP, Mg2+ and glutamate in GS. Many of the GS residues that coordinate the metal ions and interact with glutamic acid and the phosphoryl and ribosyl groups of ATP are also present in YbdK. GS amino acids that have been associated with ammonia binding have no obvious counterparts in YbdK, consistent with a substrate specificity that is different from that of GS. Ligase activity between glutamic acid and each of the twenty amino acid residues was tested on high performance liquid chromatography (HPLC) by following the hydrolysis of ATP to ADP. Catalysis was observed only with cysteine. A pyruvate kinase/lactic acid dehydrogenase coupled assay was used to rule out GS activity and to determine that YbdK exhibits gamma-GCS activity. The catalytic rate was found to be approximately 500-fold slower than that reported for authentic gamma-GCS.
About this Structure
1R8G is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.
Reference
YbdK is a carboxylate-amine ligase with a gamma-glutamyl:Cysteine ligase activity: crystal structure and enzymatic assays., Lehmann C, Doseeva V, Pullalarevu S, Krajewski W, Howard A, Herzberg O, Proteins. 2004 Aug 1;56(2):376-83. PMID:15211520
Page seeded by OCA on Thu Feb 21 14:48:15 2008
Categories: Escherichia coli | Single protein | Doseeva, V. | Herzberg, O. | Howard, A. | Krajewski, W. | Lehmann, C. | Pullalarevu, S. | S2F, Structure 2.Function Project. | Carboxylate-amine ligase | Hypothetical protein | S2f | Structural genomics | Structure 2 function project | Unknown function | Ybdk
