2ez2: Difference between revisions

Revision as of 18:15, 21 February 2008

Apo tyrosine phenol-lyase from Citrobacter freundii at pH 8.0

OverviewOverview

Tyrosine phenol-lyase, a tetrameric pyridoxal 5'-phosphate dependent enzyme, catalyzes the reversible hydrolytic cleavage of L-tyrosine to phenol and ammonium pyruvate. Here we describe the crystal structure of the Citrobacter freundii holoenzyme at 1.9 A resolution. The structure reveals a network of protein interactions with the cofactor, pyridoxal 5'-phosphate, and details of coordination of the catalytically important K+ ion. We also present the structure of the apoenzyme at 1.85 A resolution. Both structures were determined using crystals grown at pH 8.0, which is close to the pH of the maximal enzymatic activity (8.2). Comparison of the apoenzyme structure with the one previously determined at pH 6.0 reveals significant differences. The data suggest that the decrease of the enzymatic activity at pH 6.0 may be caused by conformational changes in the active site residues Tyr71, Tyr291, and Arg381 and in the monovalent cation binding residue Glu69. Moreover, at pH 8.0 we observe two different active site conformations: open, which was characterized before, and closed, which is observed for the first time in beta-eliminating lyases. In the closed conformation a significant part of the small domain undergoes an extraordinary motion of up to 12 A toward the large domain, closing the active site cleft and bringing the catalytically important Arg381 and Phe448 into the active site. The closed conformation allows rationalization of the results of previous mutational studies and suggests that the observed active site closure is critical for the course of the enzymatic reaction and for the enzyme's specificity toward its physiological substrate. Finally, the closed conformation allows us to model keto(imino)quinonoid, the key transition intermediate.

About this StructureAbout this Structure

2EZ2 is a Single protein structure of sequence from Citrobacter freundii with and as ligands. Active as Tyrosine phenol-lyase, with EC number 4.1.99.2 Full crystallographic information is available from OCA.

ReferenceReference

Structures of apo- and holo-tyrosine phenol-lyase reveal a catalytically critical closed conformation and suggest a mechanism for activation by K+ ions., Milic D, Matkovic-Calogovic D, Demidkina TV, Kulikova VV, Sinitzina NI, Antson AA, Biochemistry. 2006 Jun 20;45(24):7544-52. PMID:16768450

Page seeded by OCA on Thu Feb 21 17:15:58 2008

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OCA

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-[[Image:2ez2.gif|left|200px]]<br /><applet load="2ez2" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:2ez2.gif|left|200px]]<br /><applet load="2ez2" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="2ez2, resolution 1.850&Aring;" />
 '''Apo tyrosine phenol-lyase from Citrobacter freundii at pH 8.0'''<br />
 ==Overview==
-Tyrosine phenol-lyase, a tetrameric pyridoxal 5'-phosphate dependent, enzyme, catalyzes the reversible hydrolytic cleavage of L-tyrosine to, phenol and ammonium pyruvate. Here we describe the crystal structure of, the Citrobacter freundii holoenzyme at 1.9 A resolution. The structure, reveals a network of protein interactions with the cofactor, pyridoxal, 5'-phosphate, and details of coordination of the catalytically important, K+ ion. We also present the structure of the apoenzyme at 1.85 A, resolution. Both structures were determined using crystals grown at pH, 8.0, which is close to the pH of the maximal enzymatic activity (8.2)., Comparison of the apoenzyme structure with the one previously determined, at pH 6.0 reveals significant differences. The data suggest that the, decrease of the enzymatic activity at pH 6.0 may be caused by, conformational changes in the active site residues Tyr71, Tyr291, and, Arg381 and in the monovalent cation binding residue Glu69. Moreover, at pH, 8.0 we observe two different active site conformations: open, which was, characterized before, and closed, which is observed for the first time in, beta-eliminating lyases. In the closed conformation a significant part of, the small domain undergoes an extraordinary motion of up to 12 A toward, the large domain, closing the active site cleft and bringing the, catalytically important Arg381 and Phe448 into the active site. The closed, conformation allows rationalization of the results of previous mutational, studies and suggests that the observed active site closure is critical for, the course of the enzymatic reaction and for the enzyme's specificity, toward its physiological substrate. Finally, the closed conformation, allows us to model keto(imino)quinonoid, the key transition intermediate.
+Tyrosine phenol-lyase, a tetrameric pyridoxal 5'-phosphate dependent enzyme, catalyzes the reversible hydrolytic cleavage of L-tyrosine to phenol and ammonium pyruvate. Here we describe the crystal structure of the Citrobacter freundii holoenzyme at 1.9 A resolution. The structure reveals a network of protein interactions with the cofactor, pyridoxal 5'-phosphate, and details of coordination of the catalytically important K+ ion. We also present the structure of the apoenzyme at 1.85 A resolution. Both structures were determined using crystals grown at pH 8.0, which is close to the pH of the maximal enzymatic activity (8.2). Comparison of the apoenzyme structure with the one previously determined at pH 6.0 reveals significant differences. The data suggest that the decrease of the enzymatic activity at pH 6.0 may be caused by conformational changes in the active site residues Tyr71, Tyr291, and Arg381 and in the monovalent cation binding residue Glu69. Moreover, at pH 8.0 we observe two different active site conformations: open, which was characterized before, and closed, which is observed for the first time in beta-eliminating lyases. In the closed conformation a significant part of the small domain undergoes an extraordinary motion of up to 12 A toward the large domain, closing the active site cleft and bringing the catalytically important Arg381 and Phe448 into the active site. The closed conformation allows rationalization of the results of previous mutational studies and suggests that the observed active site closure is critical for the course of the enzymatic reaction and for the enzyme's specificity toward its physiological substrate. Finally, the closed conformation allows us to model keto(imino)quinonoid, the key transition intermediate.
 ==About this Structure==
-EZ2 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Citrobacter_freundii Citrobacter freundii] with K and PO4 as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Tyrosine_phenol-lyase Tyrosine phenol-lyase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=4.1.99.2 4.1.99.2] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=2EZ2 OCA].
+EZ2 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Citrobacter_freundii Citrobacter freundii] with <scene name='pdbligand=K:'>K</scene> and <scene name='pdbligand=PO4:'>PO4</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Tyrosine_phenol-lyase Tyrosine phenol-lyase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=4.1.99.2 4.1.99.2] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2EZ2 OCA].
 ==Reference==
@@ Line 14: / Line 14: @@
 [[Category: Single protein]]
 [[Category: Tyrosine phenol-lyase]]
-[[Category: Antson, A.A.]]
+[[Category: Antson, A A.]]
-[[Category: Demidkina, T.V.]]
+[[Category: Demidkina, T V.]]
 [[Category: Matkovic-Calogovic, D.]]
 [[Category: Milic, D.]]
@@ Line 22: / Line 22: @@
 [[Category: lyase; plp-dependent enzyme; pyridoxal-5'-phosphate; domain closure]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Wed Nov 21 10:14:28 2007''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 17:15:58 2008''