The pet operon consists of genes coding for enzymes responsible for ethanol production and consists of pyruvate dehydrogenase and alcohol dehydrogenase II from the high-performance ethanologen Zymomonas mobilis. This article describes the physiological influence of pet expression in Escherichia coli B (ATCC 11303) in terms of growth rate and overall concentrations of cell mass and catabolic end products achieved under well-defined cultivation conditions that included constant pH and carbon (energy) limitation. Glucose, mannose, and xylose were used as substrates, because they represent the principal fermentable components of lignocellulosic biomass and because fermentation of these sugars involves different metabolic pathways. Two different types of ethanologenic recombinants were used-a strain in which pet expression was via a multicopy plasmid (pLOI297) and a chromosomal integrant, strain KO11. Under the condition of sugar substrate limitation, there was no growth enhancement by pet expression with either glucose or mannose. Whereas the host strain produced exclusively lactic acid from glucose and mannose, both recombinants produced mostly ethanol. Both the plasmid-carrying strain and the per integrant exhibited slower growth compared to the host culture with glucose or mannose as fermentation substrate. With mannose, the plasmid recombinant grew appreciably slower than either the host culture or the recombinant KO11. Use of a magnesium-deficient medium produced different results with glucose since substrate and turbidometric measurements proved to be unreliable in terms of estimating overall biomass levels. At pH, 6.3, pet expression improved overall biomass yield; but at pH 7.0, the cell yields exhibited by the plasmid recombinant and the host strain were the same. E. coli B did not grow well on xylose as sole carbon source. With xylose, pet expression increased the growth rate, but had no effect on the overall biomass yield. Ln comparing our observations with the reports of others, it was concluded that the effect of pet expression on growth of E. coli is dependent on several different biochemical, physiological, genetic, and environmental factors, which largely precludes a statement of generality regarding this phenomenon.