Energy cost of galactoside transport to Escherichia coli

Abstract

Energy reserves of E. coli can be depleted by a previously reported procedure to a level such that even the 'downhill' transport of o nitrophenyl β D galactopyranoside (ONPG) is completely dependent upon the exogenous energy supply. The ONPG concentration is high externally to the cells and is low intracellular because of the action of cytoplasmic β galactosidase. In the present work, depleted cell suspensions have been infused at low, steady rates with glucose and other energy sources while measurements of transport were being made. Comparing the rate of ONPG transport with the rate of introduction of glucose under conditions where the chosen glucose infusion rate limits transport, it is found that 89 molecules of ONPG are transported per molecule of fully oxidized glucose. This transport yield is constant over a 6.5 fold range in rate of glucose addition. This constancy over a range of infusion rates implies that transport is the major cellular function under these special conditions. The yield value of 89 is in agreement with the predictions of 76 from Mitchell's chemiosmotic theory and constitutes an independent proof of its validity, since all the other proposed mechanisms of energy coupling predict much smaller yields. The lag from the start of glucose infusion into the reaction cuvette, to the extrapolated time at which a steady state rate of transport and concomitant hydrolysis are achieved, is short (1 min). Similarly, the time after the infusion is stopped until the rate of transport returns to the background rate is also short. The latter implies that the energy metabolism is directed almost entirely to transport and/or other ongoing cellular processes and not to repair or renewal of an energy independent, facilitated diffusion system.