当前位置： SCI文献检索 > BIOCHEMISTRY期刊下所有文献 > Structures of apo- and holo-tyrosine phenol-lyase reveal a catalytically critical closed conformation and suggest a mechanism for activation by K+ ions.

Structures of apo- and holo-tyrosine phenol-lyase reveal a catalytically critical closed conformation and suggest a mechanism for activation by K+ ions.

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.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Milić D,Matković-Calogović D,Demidkina TV,Kulikova VV,Sinitzina NI,Antson AA

doi

10.1021/bi0601858

subject

Has Abstract

pub_date

2006-06-20 00:00:00

pages

7544-52

issue

eissn

0006-2960

issn

1520-4995

journal_volume

pub_type

杂志文章

在线工具

Structures of apo- and holo-tyrosine phenol-lyase reveal a catalytically critical closed conformation and suggest a mechanism for activation by K+ ions.