2EZ2

Apo tyrosine phenol-lyase from Citrobacter freundii at pH 8.0

Summary for 2EZ2

• Entry DOI: 10.2210/pdb2ez2/pdb
• Related: 2EZ1
• Descriptor: Tyrosine phenol-lyase, POTASSIUM ION, PHOSPHATE ION, ... (4 entities in total)
• Functional Keywords: lyase; plp-dependent enzyme; pyridoxal-5'-phosphate; domain closure, lyase
• Biological source: Citrobacter freundii
• Total number of polymer chains: 2
• Total formula weight: 103401.71

Authors

Milic, D.,Matkovic-Calogovic, D.,Demidkina, T.V.,Antson, A.A. (deposition date: 2005-11-10, release date: 2006-07-25, Last modification date: 2024-02-14)

Primary citation

Milic, D.,Matkovic-Calogovic, D.,Demidkina, T.V.,Kulikova, V.V.,Sinitzina, N.I.,Antson, A.A.
Structures of apo- and holo-tyrosine phenol-lyase reveal a catalytically critical closed conformation and suggest a mechanism for activation by K+ ions
Biochemistry, 45:7544-7552, 2006

PubMed Abstract: 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.

PubMed: 16768450
DOI: 10.1021/bi0601858

Experimental method

X-RAY DIFFRACTION (1.85 Å)