1GGO

T453A MUTANT OF PYRUVATE, PHOSPHATE DIKINASE

1GGO の概要

• エントリーDOI: 10.2210/pdb1ggo/pdb
• 分子名称: PROTEIN (PYRUVATE, PHOSPHATE DIKINASE), SULFATE ION (3 entities in total)
• 機能のキーワード: transferase, phosphotransferase, kinase
• 由来する生物種: Clostridium symbiosum
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 96802.25

構造登録者

Li, Z.,Herzberg, O. (登録日: 2000-08-29, 公開日: 2001-01-10, 最終更新日: 2023-12-27)

主引用文献

Wei, M.,Li, Z.,Ye, D.,Herzberg, O.,Dunaway-Mariano, D.
Identification of domain-domain docking sites within Clostridium symbiosum pyruvate phosphate dikinase by amino acid replacement.
J.Biol.Chem., 275:41156-41165, 2000

PubMed Abstract: Potential domain-domain docking residues, identified from the x-ray structure of the Clostridium symbiosum apoPPDK, were replaced by site-directed mutagenesis. The steady-state and transient kinetic properties of the mutant enzymes were determined as a way of evaluating docking efficiency. PPDK mutants, in which one of two stringently conserved docking residues located on the N-terminal domain (Arg(219) and Glu(271)) was substituted, displayed largely unimpeded catalysis of the phosphoenolpyruvate partial reaction at the C-terminal domain, but significantly impaired catalysis (>10(4)) of the ATP pyrophosphorylation of His(455) at the N-terminal domain. In contrast, alanine mutants of two potential docking residues located on the N-terminal domain (Ser(262) and Lys(149)), which are not conserved among the PPDKs, exhibited essentially normal catalytic turnover. Arg(219) and Glu(271) were thus proposed to play an important role in guiding the central domain and, hence, the catalytic His(455) into position for catalysis. Substitution of central domain residues Glu(434)/Glu(437) and Thr(453), the respective docking partners of Arg(219) and Glu(271), resulted in mutants impaired in catalysis at the ATP active site. The x-ray crystal structure of the apo-T453A PPDK mutant was determined to test for possible misalignment of residues at the N-terminal domain-central domain interface that might result from loss of the Thr(453)-Glu(271) binding interaction. With the exception of the mutation site, the structure of T453A PPDK was found to be identical to that of the wild-type enzyme. It is hypothesized that the two Glu(271) interfacial binding sites that remain in the T453A PPDK mutant, Thr(453) backbone NH and Met(452) backbone NH, are sufficient to stabilize the native conformation as observed in the crystalline state but may be less effective in populating the reactive conformation in solution.

PubMed: 10995759
DOI: 10.1074/jbc.M006149200

実験手法

X-RAY DIFFRACTION (2.6 Å)