1at1: Difference between revisions

Revision as of 12:48, 21 February 2008

CRYSTAL STRUCTURES OF PHOSPHONOACETAMIDE LIGATED T AND PHOSPHONOACETAMIDE AND MALONATE LIGATED R STATES OF ASPARTATE CARBAMOYLTRANSFERASE AT 2.8-ANGSTROMS RESOLUTION AND NEUTRAL P*H

OverviewOverview

The T----R transition of the cooperative enzyme aspartate carbamoyltransferase occurs at pH 7 in single crystals without visibly cracking many of the crystals and leaving those uncracked suitable for single-crystal X-ray analysis. To promote the T----R transition, we employ the competitive inhibitors of carbamoyl phosphate and aspartate, which are phosphonoacetamide (PAM) and malonate, respectively. In response to PAM binding to the T-state crystals, residues Thr 53-Thr 55 and Pro 266-Pro 268 move to their R-state positions to bind to the phosphonate and amino group of PAM. These changes induce a conformation that can bind tightly the aspartate analogue malonate, which thereby effects the allosteric transition. We prove this by showing that PAM-ligated T-state crystals (Tpam), space group P321 (a = 122.2 A, c = 142.2 A), when transferred to a solution containing 20 mM PAM and 8 mM malonate at pH 7, isomerize to R-state crystals (Rpam,mal,soak), space group also P321 (a = 122.2 A, c = 156.4 A). The R-state structure in which the T----R transition occurs within the crystal at pH 7 compares very well (rms = 0.19 A for all atoms) with an R-state structure determined at pH 7 in which the crystals were initially grown in a solution of PAM and malonate at pH 5.9 and subsequently transferred to a buffer containing the ligands at pH 7 (Rpam,mal,crys). In fact, both of the PAM and malonate ligated R-state structures are very similar to both the carbamoyl phosphate and succinate or the N-(phosphonoacetyl)-L-aspartate ligated structures, even though the R-state structures reported here were determined at pH 7. Crystallographic residuals refined to 0.16-0.18 at 2.8-A resolution for the three structures.

About this StructureAbout this Structure

1AT1 is a Protein complex structure of sequences from Escherichia coli with , and as ligands. Active as Aspartate carbamoyltransferase, with EC number 2.1.3.2 Full crystallographic information is available from OCA.

ReferenceReference

Crystal structures of phosphonoacetamide ligated T and phosphonoacetamide and malonate ligated R states of aspartate carbamoyltransferase at 2.8-A resolution and neutral pH., Gouaux JE, Lipscomb WN, Biochemistry. 1990 Jan 16;29(2):389-402. PMID:2405902

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-[[Image:1at1.gif|left|200px]]<br /><applet load="1at1" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:1at1.gif|left|200px]]<br /><applet load="1at1" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="1at1, resolution 2.8&Aring;" />
 '''CRYSTAL STRUCTURES OF PHOSPHONOACETAMIDE LIGATED T AND PHOSPHONOACETAMIDE AND MALONATE LIGATED R STATES OF ASPARTATE CARBAMOYLTRANSFERASE AT 2.8-ANGSTROMS RESOLUTION AND NEUTRAL P*H'''<br />
 ==Overview==
-The T----R transition of the cooperative enzyme aspartate, carbamoyltransferase occurs at pH 7 in single crystals without visibly, cracking many of the crystals and leaving those uncracked suitable for, single-crystal X-ray analysis. To promote the T----R transition, we employ, the competitive inhibitors of carbamoyl phosphate and aspartate, which are, phosphonoacetamide (PAM) and malonate, respectively. In response to PAM, binding to the T-state crystals, residues Thr 53-Thr 55 and Pro 266-Pro, 268 move to their R-state positions to bind to the phosphonate and amino, group of PAM. These changes induce a conformation that can bind tightly, the aspartate analogue malonate, which thereby effects the allosteric, transition. We prove this by showing that PAM-ligated T-state crystals, (Tpam), space group P321 (a = 122.2 A, c = 142.2 A), when transferred to a, solution containing 20 mM PAM and 8 mM malonate at pH 7, isomerize to, R-state crystals (Rpam,mal,soak), space group also P321 (a = 122.2 A, c =, 156.4 A). The R-state structure in which the T----R transition occurs, within the crystal at pH 7 compares very well (rms = 0.19 A for all atoms), with an R-state structure determined at pH 7 in which the crystals were, initially grown in a solution of PAM and malonate at pH 5.9 and, subsequently transferred to a buffer containing the ligands at pH 7, (Rpam,mal,crys). In fact, both of the PAM and malonate ligated R-state, structures are very similar to both the carbamoyl phosphate and succinate, or the N-(phosphonoacetyl)-L-aspartate ligated structures, even though the, R-state structures reported here were determined at pH 7. Crystallographic, residuals refined to 0.16-0.18 at 2.8-A resolution for the three, structures.
+The T----R transition of the cooperative enzyme aspartate carbamoyltransferase occurs at pH 7 in single crystals without visibly cracking many of the crystals and leaving those uncracked suitable for single-crystal X-ray analysis. To promote the T----R transition, we employ the competitive inhibitors of carbamoyl phosphate and aspartate, which are phosphonoacetamide (PAM) and malonate, respectively. In response to PAM binding to the T-state crystals, residues Thr 53-Thr 55 and Pro 266-Pro 268 move to their R-state positions to bind to the phosphonate and amino group of PAM. These changes induce a conformation that can bind tightly the aspartate analogue malonate, which thereby effects the allosteric transition. We prove this by showing that PAM-ligated T-state crystals (Tpam), space group P321 (a = 122.2 A, c = 142.2 A), when transferred to a solution containing 20 mM PAM and 8 mM malonate at pH 7, isomerize to R-state crystals (Rpam,mal,soak), space group also P321 (a = 122.2 A, c = 156.4 A). The R-state structure in which the T----R transition occurs within the crystal at pH 7 compares very well (rms = 0.19 A for all atoms) with an R-state structure determined at pH 7 in which the crystals were initially grown in a solution of PAM and malonate at pH 5.9 and subsequently transferred to a buffer containing the ligands at pH 7 (Rpam,mal,crys). In fact, both of the PAM and malonate ligated R-state structures are very similar to both the carbamoyl phosphate and succinate or the N-(phosphonoacetyl)-L-aspartate ligated structures, even though the R-state structures reported here were determined at pH 7. Crystallographic residuals refined to 0.16-0.18 at 2.8-A resolution for the three structures.
 ==About this Structure==
-AT1 is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with MAL, ZN and PCT as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Aspartate_carbamoyltransferase Aspartate carbamoyltransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.1.3.2 2.1.3.2] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1AT1 OCA].
+AT1 is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with <scene name='pdbligand=MAL:'>MAL</scene>, <scene name='pdbligand=ZN:'>ZN</scene> and <scene name='pdbligand=PCT:'>PCT</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Aspartate_carbamoyltransferase Aspartate carbamoyltransferase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.1.3.2 2.1.3.2] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1AT1 OCA].
 ==Reference==
@@ Line 14: / Line 14: @@
 [[Category: Escherichia coli]]
 [[Category: Protein complex]]
-[[Category: Gouaux, J.E.]]
+[[Category: Gouaux, J E.]]
-[[Category: Lipscomb, W.N.]]
+[[Category: Lipscomb, W N.]]
 [[Category: MAL]]
 [[Category: PCT]]
@@ Line 22: / Line 22: @@
 [[Category: transferase (carbamoyl-p]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 11:07:04 2007''
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