Increased Phenylalanine Production by Growing and Nongrowing Escherichia coli Strain CWML2

Abstract

Chemostat selection at low dilution rate in glycerol-limited minimal medium was previously employed to isolate the mutant Escherichia coli strain CWML2 which exhibits shorter lag phases, decreased acetate production, and higher specific growth rates and biomass yields in batch culture (Weikert, C.; Sauer, U.; Bailey, J. E. Microbiology 1997, 143, 1567-1574). In this study, CWML2 was analyzed for its biochemical production capabilities in batch culture and under nongrowing conditions. Both CWML2 and MG1655 were transformed with plasmid pSY130-14, which encodes feedback resistant mutants of the enzymes chorismate mutase P-prephenate synthase and 3-hydroxy-d-arabinoheptulosonate-7-phosphate dehydratase, to enable phenylalanine production. In batch culture, transformed CWML2 produced twice as much phenylalanine as did MG1655:pSY130-14. In contrast to the reference strain, substantial growth-independent production of phenylalanine was calculated for CWML2:pSY130-14 by using Luedeking-Piret kinetic analysis. Over a period of 30 h, nongrowing cells of CWML2:pSY130-14 exhibited a 2.5-fold higher specific phenylalanine production rate. The apparent capability of E. coli CWML2 to partly uncouple metabolic activity from growth suggests a potentially general advantage of this class of modified hosts for production of biochemicals.

Production of l-phenylalanine from glycerol by a recombinant Escherichia coli

“…For example, for two recombinant E. coli strains, the yields on glucose were 0.031 and 0.057 g g ¡1 , respectively [31]. A high product yield on glycerol is a reXection of a high expression level of L-phenylalanine dehydrogenase in the strain used and the fact that the three-carbon molecule of glycerol is easier to metabolize by glycolysis than a six-carbon sugar is.…”

“…The production of L-phenylalanine by various recombinant E. coli strains using glucose or sucrose as carbon sources has been discussed extensively in the literature [2,8,9,14,19,26,31,32], but reports on the use of glycerol for this fermentation are nonexistent.…”

Process control for enhanced L‐phenylalanine production using different recombinant Escherichia coli strains

“…Beyond several approaches using isolated enzymes (Asano et al, 1987;Hummel et al, 1987;Nakamichi et al, 1989;Ziehr et al, 1987), resting cells , or immobilized cells (patent Wood and Calton, US-A-4728611, 1988), fermentation process development mainly focused on E. coli (Backman et al, 1989;Choi and Tribe, 1982;Gerigk et al, 2002;Grinter, 1998;Hwang et al, 1985;Park and Rogers, 1988;Maass et al, 2001 (in press, Bioprocess and Biosystems Engineering); Sugimoto et al, 1990;Weikert et al, 1998) leading to several patent applications (Kim et al, US-A-5304475, 1994;Lee et al, US-A-5008190, 1991;Lim et al, US-A-5409830, 1995;Sprenger et al, WO-98/18937, 1998).…”

Enhanced pilot-scale fed-batch L-phenylalanine production with recombinant Escherichia coli by fully integrated reactive extraction

“…Co-expression of enzymes was carried out with Escherichia coli or Candida boidinii [6] and homologue bioconversion from phenylpyruvate to L-Phe by resting [7], immobilised [8], or growing E. coli [9,10] or Corynebacterium glutamicum cells [11,12]. Fermentation processes have been developed and optimised for mutants or genetically engineered from strains of C. glutamicum [35,36,37], Brevibacterium lactofermentum [38], Bacillus sublitis [39] or B. flavum [40] and especially of E. coli [13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34].…”