Quantitative aspects of glucose metabolism by Escherichia coli B/r, grown in the presence of pyrroloquinoline quinone

Abstract

Escherichia coli B/r was grown in chemostat cultures under various limitations with glucose as carbon source. Since E. coli only synthesized the glucose dehydrogenase (GDH) apo-enzyme and not the appropriate cofactor, pyrroloquinoline quinone (POO), no gluconate production could be observed. However, when cell-saturating amounts of POO (nmol to μmol range) were pulsed into steady state glucose-excess cultures of E. coli, the organisms responded with an instantaneous formation of gluconate and an increased oxygen consumption rate. This showed that reconstitution of GDH in situ was possible.

Hence, in order to examine the influence on glucose metabolism of an active GDH, E. coli was grown aerobically in chemostat cultures under various limitations in the presence of POO. It was found that the presence of POO indeed had a sizable effect: at pH 5.5 under phosphate-or sulphate-limited conditions more than 60% of the glucose consumed was converted to gluconate, which resulted in steady state gluconate concentrations up to 80 mmolli. The specific rate of gluconate production (0.3–7.6 mmol·h⁻¹·(g dry wt cells)⁻) was dependent on the growth rate and the nature of the limitation. The production rate of other overflow metabolites such as acetate, pyruvate, and 2-oxoglutarate, was only slightly altered in the presence of POO. The fact that the cells were now able to use an active GDH apparently did not affect apo-enzyme synthesis.