1os1
From Proteopedia
(New page: 200px<br /><applet load="1os1" size="450" color="white" frame="true" align="right" spinBox="true" caption="1os1, resolution 1.8Å" /> '''Structure of Phosphoe...) |
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| - | [[Image:1os1.jpg|left|200px]]<br /><applet load="1os1" size=" | + | [[Image:1os1.jpg|left|200px]]<br /><applet load="1os1" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1os1, resolution 1.8Å" /> | caption="1os1, resolution 1.8Å" /> | ||
'''Structure of Phosphoenolpyruvate Carboxykinase complexed with ATP,Mg, Ca and pyruvate.'''<br /> | '''Structure of Phosphoenolpyruvate Carboxykinase complexed with ATP,Mg, Ca and pyruvate.'''<br /> | ||
==Overview== | ==Overview== | ||
| - | The 1.8-A resolution structure of the ATP-Mg(2+)-Ca(2+)-pyruvate quinary | + | The 1.8-A resolution structure of the ATP-Mg(2+)-Ca(2+)-pyruvate quinary complex of Escherichia coli phosphoenolpyruvate carboxykinase (PCK) is isomorphous to the published complex ATP-Mg(2+)-Mn(2+)-pyruvate-PCK, except for the Ca(2+) and Mn(2+) binding sites. Ca(2+) was formerly implicated as a possible allosteric regulator of PCK, binding at the active site and at a surface activating site (Glu508 and Glu511). This report found that Ca(2+) bound only at the active site, indicating that there is likely no surface allosteric site. (45)Ca(2+) bound to PCK with a K(d) of 85 micro M and n of 0.92. Glu508Gln Glu511Gln mutant PCK had normal activation by Ca(2+). Separate roles of Mg(2+), which binds the nucleotide, and Ca(2+), which bridges the nucleotide and the anionic substrate, are implied, and the catalytic mechanism of PCK is better explained by studies of the Ca(2+)-bound structure. Partial trypsin digestion abolishes Ca(2+) activation (desensitizes PCK). N-terminal sequencing identified sensitive sites, i.e., Arg2 and Arg396. Arg2Ser, Arg396Ser, and Arg2Ser Arg396Ser (double mutant) PCKs altered the kinetics of desensitization. C-terminal residues 397 to 540 were removed by trypsin when wild-type PCK was completely desensitized. Phe409 and Phe413 interact with residues in the Ca(2+) binding site, probably stabilizing the C terminus. Phe409Ala, DeltaPhe409, Phe413Ala, Delta397-521 (deletion of residues 397 to 521), Arg396(TAA) (stop codon), and Asp269Glu (Ca(2+) site) mutations failed to desensitize PCK and, with the exception of Phe409Ala, appeared to have defects in the synthesis or assembly of PCK, suggesting that the structure of the C-terminal domain is important in these processes. |
==About this Structure== | ==About this Structure== | ||
| - | 1OS1 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with MG, CA, ATP and PYR as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Phosphoenolpyruvate_carboxykinase_(ATP) Phosphoenolpyruvate carboxykinase (ATP)], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=4.1.1.49 4.1.1.49] Full crystallographic information is available from [http:// | + | 1OS1 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with <scene name='pdbligand=MG:'>MG</scene>, <scene name='pdbligand=CA:'>CA</scene>, <scene name='pdbligand=ATP:'>ATP</scene> and <scene name='pdbligand=PYR:'>PYR</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Phosphoenolpyruvate_carboxykinase_(ATP) Phosphoenolpyruvate carboxykinase (ATP)], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=4.1.1.49 4.1.1.49] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1OS1 OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Phosphoenolpyruvate carboxykinase (ATP)]] | [[Category: Phosphoenolpyruvate carboxykinase (ATP)]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Delbaere, L | + | [[Category: Delbaere, L T.]] |
[[Category: Goldie, D.]] | [[Category: Goldie, D.]] | ||
[[Category: Goldie, H.]] | [[Category: Goldie, H.]] | ||
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[[Category: phosphotransfer]] | [[Category: phosphotransfer]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 14:20:56 2008'' |
Revision as of 12:20, 21 February 2008
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Structure of Phosphoenolpyruvate Carboxykinase complexed with ATP,Mg, Ca and pyruvate.
Overview
The 1.8-A resolution structure of the ATP-Mg(2+)-Ca(2+)-pyruvate quinary complex of Escherichia coli phosphoenolpyruvate carboxykinase (PCK) is isomorphous to the published complex ATP-Mg(2+)-Mn(2+)-pyruvate-PCK, except for the Ca(2+) and Mn(2+) binding sites. Ca(2+) was formerly implicated as a possible allosteric regulator of PCK, binding at the active site and at a surface activating site (Glu508 and Glu511). This report found that Ca(2+) bound only at the active site, indicating that there is likely no surface allosteric site. (45)Ca(2+) bound to PCK with a K(d) of 85 micro M and n of 0.92. Glu508Gln Glu511Gln mutant PCK had normal activation by Ca(2+). Separate roles of Mg(2+), which binds the nucleotide, and Ca(2+), which bridges the nucleotide and the anionic substrate, are implied, and the catalytic mechanism of PCK is better explained by studies of the Ca(2+)-bound structure. Partial trypsin digestion abolishes Ca(2+) activation (desensitizes PCK). N-terminal sequencing identified sensitive sites, i.e., Arg2 and Arg396. Arg2Ser, Arg396Ser, and Arg2Ser Arg396Ser (double mutant) PCKs altered the kinetics of desensitization. C-terminal residues 397 to 540 were removed by trypsin when wild-type PCK was completely desensitized. Phe409 and Phe413 interact with residues in the Ca(2+) binding site, probably stabilizing the C terminus. Phe409Ala, DeltaPhe409, Phe413Ala, Delta397-521 (deletion of residues 397 to 521), Arg396(TAA) (stop codon), and Asp269Glu (Ca(2+) site) mutations failed to desensitize PCK and, with the exception of Phe409Ala, appeared to have defects in the synthesis or assembly of PCK, suggesting that the structure of the C-terminal domain is important in these processes.
About this Structure
1OS1 is a Single protein structure of sequence from Escherichia coli with , , and as ligands. Active as Phosphoenolpyruvate carboxykinase (ATP), with EC number 4.1.1.49 Full crystallographic information is available from OCA.
Reference
Mechanisms of activation of phosphoenolpyruvate carboxykinase from Escherichia coli by Ca2+ and of desensitization by trypsin., Sudom A, Walters R, Pastushok L, Goldie D, Prasad L, Delbaere LT, Goldie H, J Bacteriol. 2003 Jul;185(14):4233-42. PMID:12837799
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