2z2a

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(New page: 200px<br /><applet load="2z2a" size="350" color="white" frame="true" align="right" spinBox="true" caption="2z2a, resolution 1.87&Aring;" /> '''Thr109Gly dihydrooro...)
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==Overview==
==Overview==
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Dihydroorotase (DHOase) catalyzes the reversible cyclization of, N-carbamyl-l-aspartate (CA-asp) to l-dihydroorotate (DHO) in the de novo, biosynthesis of pyrimidine nucleotides. Two different conformations of the, surface loop (residues 105-115) were found in the dimeric Escherichia coli, DHOase crystallized in the presence of DHO (PDB code 1XGE). The loop, asymmetry reflected that of the active site contents of the two subunits:, the product, DHO, was bound in the active site of one subunit and the, substrate, CA-asp, in the active site of the other. In the substrate-, (CA-asp-) bound subunit, the surface loop reaches in toward the active, site and makes hydrogen bonds with the bound CA-asp via two threonine, residues (Thr109 and Thr110), whereas the loop forms part of the surface, of the protein in the product- (DHO-) bound subunit. To investigate the, relationship between the structural states of this loop and the catalytic, mechanism of the enzyme, a series of mutant DHOases including deletion of, the flexible loop were generated and characterized kinetically and, structurally. Disruption of the hydrogen bonds between the surface loop, and the substrate results in significant loss of catalytic activity., Furthermore, structures of these mutants with low catalytic activity have, no interpretable electron density for parts of the flexible loop. The, structure of the mutant (Delta107-116), in which the flexible loop is, deleted, shows only small differences in positions of other substrate, binding residues and in the binuclear zinc center compared with the native, structure, yet the enzyme has negligible activity. The kinetic and, structural analyses suggest that Thr109 and Thr110 in the flexible loop, provide productive binding of substrate and stabilize the transition-state, intermediate, thereby increasing catalytic activity.
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Dihydroorotase (DHOase) catalyzes the reversible cyclization of N-carbamyl-l-aspartate (CA-asp) to l-dihydroorotate (DHO) in the de novo biosynthesis of pyrimidine nucleotides. Two different conformations of the surface loop (residues 105-115) were found in the dimeric Escherichia coli DHOase crystallized in the presence of DHO (PDB code 1XGE). The loop asymmetry reflected that of the active site contents of the two subunits: the product, DHO, was bound in the active site of one subunit and the substrate, CA-asp, in the active site of the other. In the substrate- (CA-asp-) bound subunit, the surface loop reaches in toward the active site and makes hydrogen bonds with the bound CA-asp via two threonine residues (Thr109 and Thr110), whereas the loop forms part of the surface of the protein in the product- (DHO-) bound subunit. To investigate the relationship between the structural states of this loop and the catalytic mechanism of the enzyme, a series of mutant DHOases including deletion of the flexible loop were generated and characterized kinetically and structurally. Disruption of the hydrogen bonds between the surface loop and the substrate results in significant loss of catalytic activity. Furthermore, structures of these mutants with low catalytic activity have no interpretable electron density for parts of the flexible loop. The structure of the mutant (Delta107-116), in which the flexible loop is deleted, shows only small differences in positions of other substrate binding residues and in the binuclear zinc center compared with the native structure, yet the enzyme has negligible activity. The kinetic and structural analyses suggest that Thr109 and Thr110 in the flexible loop provide productive binding of substrate and stabilize the transition-state intermediate, thereby increasing catalytic activity.
==About this Structure==
==About this Structure==
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==Reference==
==Reference==
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Kinetic and Structural Analysis of Mutant Escherichia coli Dihydroorotases: A Flexible Loop Stabilizes the Transition State(,)., Lee M, Maher MJ, Christopherson RI, Guss JM, Biochemistry. 2007 Sep 18;46(37):10538-50. Epub 2007 Aug 21. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17711307 17711307]
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Kinetic and structural analysis of mutant Escherichia coli dihydroorotases: a flexible loop stabilizes the transition state., Lee M, Maher MJ, Christopherson RI, Guss JM, Biochemistry. 2007 Sep 18;46(37):10538-50. Epub 2007 Aug 21. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17711307 17711307]
[[Category: Dihydroorotase]]
[[Category: Dihydroorotase]]
[[Category: Escherichia coli]]
[[Category: Escherichia coli]]
[[Category: Single protein]]
[[Category: Single protein]]
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[[Category: Guss, J.M.]]
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[[Category: Guss, J M.]]
[[Category: Lee, M.]]
[[Category: Lee, M.]]
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[[Category: Maher, M.J.]]
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[[Category: Maher, M J.]]
[[Category: DOR]]
[[Category: DOR]]
[[Category: NCD]]
[[Category: NCD]]
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[[Category: tim barrel]]
[[Category: tim barrel]]
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Wed Jan 23 13:05:54 2008''
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 18:59:40 2008''

Revision as of 16:59, 21 February 2008

Image:2z2a.gif

2z2a, resolution 1.87Å

Drag the structure with the mouse to rotate

Thr109Gly dihydroorotase from E. coli

Overview

Dihydroorotase (DHOase) catalyzes the reversible cyclization of N-carbamyl-l-aspartate (CA-asp) to l-dihydroorotate (DHO) in the de novo biosynthesis of pyrimidine nucleotides. Two different conformations of the surface loop (residues 105-115) were found in the dimeric Escherichia coli DHOase crystallized in the presence of DHO (PDB code 1XGE). The loop asymmetry reflected that of the active site contents of the two subunits: the product, DHO, was bound in the active site of one subunit and the substrate, CA-asp, in the active site of the other. In the substrate- (CA-asp-) bound subunit, the surface loop reaches in toward the active site and makes hydrogen bonds with the bound CA-asp via two threonine residues (Thr109 and Thr110), whereas the loop forms part of the surface of the protein in the product- (DHO-) bound subunit. To investigate the relationship between the structural states of this loop and the catalytic mechanism of the enzyme, a series of mutant DHOases including deletion of the flexible loop were generated and characterized kinetically and structurally. Disruption of the hydrogen bonds between the surface loop and the substrate results in significant loss of catalytic activity. Furthermore, structures of these mutants with low catalytic activity have no interpretable electron density for parts of the flexible loop. The structure of the mutant (Delta107-116), in which the flexible loop is deleted, shows only small differences in positions of other substrate binding residues and in the binuclear zinc center compared with the native structure, yet the enzyme has negligible activity. The kinetic and structural analyses suggest that Thr109 and Thr110 in the flexible loop provide productive binding of substrate and stabilize the transition-state intermediate, thereby increasing catalytic activity.

About this Structure

2Z2A is a Single protein structure of sequence from Escherichia coli with , and as ligands. Active as Dihydroorotase, with EC number 3.5.2.3 Full crystallographic information is available from OCA.

Reference

Kinetic and structural analysis of mutant Escherichia coli dihydroorotases: a flexible loop stabilizes the transition state., Lee M, Maher MJ, Christopherson RI, Guss JM, Biochemistry. 2007 Sep 18;46(37):10538-50. Epub 2007 Aug 21. PMID:17711307

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