2DIK

R337A MUTANT OF PYRUVATE PHOSPHATE DIKINASE

「1BUK」から置き換えられました

2DIK の概要

• エントリーDOI: 10.2210/pdb2dik/pdb
• 分子名称: PROTEIN (PYRUVATE PHOSPHATE DIKINASE), SULFATE ION (3 entities in total)
• 機能のキーワード: phosphotransferase, kinase, transferase
• 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: 96760.18

構造登録者

Huang, K.,Herzberg, O. (登録日: 1998-09-03, 公開日: 1999-09-13, 最終更新日: 2023-08-23)

主引用文献

McGuire, M.,Huang, K.,Kapadia, G.,Herzberg, O.,Dunaway-Mariano, D.
Location of the phosphate binding site within Clostridium symbiosum pyruvate phosphate dikinase.
Biochemistry, 37:13463-13474, 1998

PubMed Abstract: Pyruvate phosphate dikinase (PPDK) catalyzes the interconversion of ATP, Pi, and pyruvate with AMP, PPi, and PEP in three partial reactions: (1) E + ATP --> E.ATP --> E-PP.AMP, (2) E-PP.AMP + Pi --> E-PP.AMP.Pi --> E-P.AMP.PPi, and (3) E-P + pyruvate --> E-P.pyruvate --> E.PEP. The Clostridium symbiosum PPDK structure consists of N-terminal, central, and C-terminal domains. The N-terminal and central domains catalyze partial reactions 1 and 2 whereas the C-terminal and central domains catalyze partial reaction 3. The goal of the present work is to determine where on the N-terminal domain catalysis of partial reactions 1 and 2 occurs and, in particular, where the Pi binding site is located. Computer modeling studies implicated Arg337 as a key residue for Pi binding. This role was tested by site-directed mutagenesis. The R337A PPDK was shown to be impaired in catalysis of the forward (kcat 300-fold lower) and reverse (kcat 30-fold lower) full reactions. Time courses for the single turnover reactions were measured to show that catalysis of partial reaction 1 is 5-fold slower in the mutant, catalysis of the second partial reaction is 140-fold slower in the mutant, and catalysis of the third partial reaction is unaffected. With the exception of the mutation site, the crystal structure of the R337A PPDK closely resembles the structure of the wild-type protein. Thus, the altered kinetic properties observed for this mutant are attributed solely to the elimination of the interaction between substrate and the guanidinium group of the Arg337 side chain. On the basis of these findings we propose that the Pi binding site is located within the crevice of the PPDK N-terminal domain, at a site that is flanked by the ATP beta-P and the Mg2+ cofactor.

PubMed: 9753432
DOI: 10.1021/bi980920i

実験手法

X-RAY DIFFRACTION (2.5 Å)