1c30: Difference between revisions

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CRYSTAL STRUCTURE OF CARBAMOYL PHOSPHATE SYNTHETASE: SMALL SUBUNIT MUTATION C269S

OverviewOverview

Carbamoyl phosphate synthetase (CPS) plays a key role in both arginine and pyrimidine biosynthesis by catalyzing the production of carbamoyl phosphate. The enzyme from Escherichi coli consists of two polypeptide chains referred to as the small and large subunits. On the basis of both amino acid sequence analyses and X-ray structural studies, it is known that the small subunit belongs to the Triad or Type I class of amidotransferases, all of which contain a cysteine-histidine (Cys269 and His353) couple required for activity. The hydrolysis of glutamine by the small subunit has been proposed to occur via two tetrahedral intermediates and a glutamyl-thioester moiety. Here, we describe the three-dimensional structures of the C269S/glutamine and CPS/glutamate gamma-semialdehyde complexes, which serve as mimics for the Michaelis complex and the tetrahedral intermediates, respectively. In conjunction with the previously solved glutamyl-thioester intermediate complex, the stereochemical course of glutamine hydrolysis in CPS has been outlined. Specifically, attack by the thiolate of Cys269 occurs at the Si face of the carboxamide group of the glutamine substrate leading to a tetrahedral intermediate with an S-configuration. Both the backbone amide groups of Gly241 and Leu270, and O(gamma) of Ser47 play key roles in stabilizing the developing oxyanion. Collapse of the tetrahedral intermediate leads to formation of the glutamyl-thioester intermediate, which is subsequently attacked at the Si face by an activated water molecule positioned near His353. The results described here serve as a paradigm for other members of the Triad class of amidotranferases.

About this StructureAbout this Structure

1C30 is a Protein complex structure of sequences from [1] with , , , , , and as ligands. Active as Carbamoyl-phosphate synthase (glutamine-hydrolyzing), with EC number 6.3.5.5 Full crystallographic information is available from OCA.

ReferenceReference

The small subunit of carbamoyl phosphate synthetase: snapshots along the reaction pathway., Thoden JB, Huang X, Raushel FM, Holden HM, Biochemistry. 1999 Dec 7;38(49):16158-66. PMID:10587438 [[Category: ]]

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-[[Image:1c30.gif|left|200px]]<br /><applet load="1c30" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:1c30.gif|left|200px]]<br /><applet load="1c30" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="1c30, resolution 2.00&Aring;" />
 '''CRYSTAL STRUCTURE OF CARBAMOYL PHOSPHATE SYNTHETASE: SMALL SUBUNIT MUTATION C269S'''<br />
 ==Overview==
-Carbamoyl phosphate synthetase (CPS) plays a key role in both arginine and, pyrimidine biosynthesis by catalyzing the production of carbamoyl, phosphate. The enzyme from Escherichi coli consists of two polypeptide, chains referred to as the small and large subunits. On the basis of both, amino acid sequence analyses and X-ray structural studies, it is known, that the small subunit belongs to the Triad or Type I class of, amidotransferases, all of which contain a cysteine-histidine (Cys269 and, His353) couple required for activity. The hydrolysis of glutamine by the, small subunit has been proposed to occur via two tetrahedral intermediates, and a glutamyl-thioester moiety. Here, we describe the three-dimensional, structures of the C269S/glutamine and CPS/glutamate gamma-semialdehyde, complexes, which serve as mimics for the Michaelis complex and the, tetrahedral intermediates, respectively. In conjunction with the, previously solved glutamyl-thioester intermediate complex, the, stereochemical course of glutamine hydrolysis in CPS has been outlined., Specifically, attack by the thiolate of Cys269 occurs at the Si face of, the carboxamide group of the glutamine substrate leading to a tetrahedral, intermediate with an S-configuration. Both the backbone amide groups of, Gly241 and Leu270, and O(gamma) of Ser47 play key roles in stabilizing the, developing oxyanion. Collapse of the tetrahedral intermediate leads to, formation of the glutamyl-thioester intermediate, which is subsequently, attacked at the Si face by an activated water molecule positioned near, His353. The results described here serve as a paradigm for other members, of the Triad class of amidotranferases.
+Carbamoyl phosphate synthetase (CPS) plays a key role in both arginine and pyrimidine biosynthesis by catalyzing the production of carbamoyl phosphate. The enzyme from Escherichi coli consists of two polypeptide chains referred to as the small and large subunits. On the basis of both amino acid sequence analyses and X-ray structural studies, it is known that the small subunit belongs to the Triad or Type I class of amidotransferases, all of which contain a cysteine-histidine (Cys269 and His353) couple required for activity. The hydrolysis of glutamine by the small subunit has been proposed to occur via two tetrahedral intermediates and a glutamyl-thioester moiety. Here, we describe the three-dimensional structures of the C269S/glutamine and CPS/glutamate gamma-semialdehyde complexes, which serve as mimics for the Michaelis complex and the tetrahedral intermediates, respectively. In conjunction with the previously solved glutamyl-thioester intermediate complex, the stereochemical course of glutamine hydrolysis in CPS has been outlined. Specifically, attack by the thiolate of Cys269 occurs at the Si face of the carboxamide group of the glutamine substrate leading to a tetrahedral intermediate with an S-configuration. Both the backbone amide groups of Gly241 and Leu270, and O(gamma) of Ser47 play key roles in stabilizing the developing oxyanion. Collapse of the tetrahedral intermediate leads to formation of the glutamyl-thioester intermediate, which is subsequently attacked at the Si face by an activated water molecule positioned near His353. The results described here serve as a paradigm for other members of the Triad class of amidotranferases.
 ==About this Structure==
-C30 is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/ ] with MN, K, CL, PO4, ORN, ADP and NET as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Carbamoyl-phosphate_synthase_(glutamine-hydrolyzing) Carbamoyl-phosphate synthase (glutamine-hydrolyzing)], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=6.3.5.5 6.3.5.5] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1C30 OCA].
+C30 is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/ ] with <scene name='pdbligand=MN:'>MN</scene>, <scene name='pdbligand=K:'>K</scene>, <scene name='pdbligand=CL:'>CL</scene>, <scene name='pdbligand=PO4:'>PO4</scene>, <scene name='pdbligand=ORN:'>ORN</scene>, <scene name='pdbligand=ADP:'>ADP</scene> and <scene name='pdbligand=NET:'>NET</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Carbamoyl-phosphate_synthase_(glutamine-hydrolyzing) Carbamoyl-phosphate synthase (glutamine-hydrolyzing)], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=6.3.5.5 6.3.5.5] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1C30 OCA].
 ==Reference==
@@ Line 14: / Line 14: @@
 [[Category: Carbamoyl-phosphate synthase (glutamine-hydrolyzing)]]
 [[Category: Protein complex]]
-[[Category: Holden, H.M.]]
+[[Category: Holden, H M.]]
 [[Category: Huang, X.]]
-[[Category: Raushel, F.M.]]
+[[Category: Raushel, F M.]]
-[[Category: Thoden, J.B.]]
+[[Category: Thoden, J B.]]
 [[Category: ADP]]
 [[Category: CL]]
@@ Line 29: / Line 29: @@
 [[Category: substrate channeling]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 12:07:29 2007''
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