Properties of an Escherichia coli mutant deficient in phosphoenolpyruvate carboxylase catalytic activity

Abstract

A mutant Escherichia coli (Ppcc') which was unable to grow on glucose as a sole carbon source was isolated. This mutant had very low levels of phosphoenolpyruvate carboxylase activity (approximately 5% of the wild type). Goat immunoglobulin G prepared against wild-type phosphoenolypyruvate carboxylase cross-reacted with the Ppccenzyme. The amount of enzyme protein in the mutant cells was similar to that found in wild-type cells, but it had greatly diminished specific activity. The catalytically less active mutant enzyme retained the ability to interact with fructose 1,6-bisphosphate, but did not exhibit stabilization of the tetrameric form by aspartate. The pl of the mutant protein was lower (4.9) than that of the wild-type protein (5.1). After electrophoresis and immunoblotting of the partially purified protein, several immunostaining bands were seen in addition to the main enzyme band. A novel method for showing that these bands represented proteolytic fragments of phosphoenolpyruvate carboxylase was developed.

Fundamental Escherichia coli biochemical pathways for biomass and energy production: Identification of reactions

“…However, the TCA cycle cannot, under steady-state conditions, supply all of the metabolite required for biomass production. This prediction is consistent with experimental data that has shown E. coli is unable to grow on glucose minimal media without the PEP carboxylase enzyme (Coomes et al, 1985;Kornberg, 1966aKornberg, , 1966b.…”

Site‐directed mutagenesis of Flaveria trinervia phosphoenolpyruvate carboxylase: Arg450 and Arg767 are essential for catalytic activity and Lys829 affects substrate binding

“…After 24 h the absorbance increased to 1.54 and 1.22, respectively. It has been observed that E. coli cells containing low PEPC activity take longer to adapt to and grow poorly in minimal medium [20,25]. The decreased cell growth indicated that Gly 829 PEPC might have lower activity than wild-type enzyme.…”

Multiobjective strain design: A framework for modular cell engineering

“…Likewise, deletion of carbon dioxide and water transport and exchange reactions caused compatibility loss across all designs. Pyruvate carboxylase (PPC) is an important reaction to channel carbon flux through the Krebs cycle (56), and hence, deletion of PPC reduces compatibility in most modular cell designs with an average CL of 43%.…”