7 ± 1.29 min versus 31.8 ± 1.29 min, RG7112 P < 0.01). The addition of 10 mM glucose at OD600 of 1 increased the growth rate of the wild-type but had only a minor effect on that of the mutant (Fig.

1). 60 min after glucose addition, glucose was depleted from the medium down to 0.3 mM by the wild-type, while still 3 mM of glucose were left in the culture of the mutant (Fig. 1). Despite increased growth and glucose consumption rates in the wild-type culture, acetate production was only slightly enhanced compared to the mutant, in line with previous findings [24]. No lactate was excreted under these conditions at any time point sampled, confirming the aerobic growth conditions. Acidification of the medium upon glucose metabolism was prevented by HEPES-buffering, which allowed maintaining the pH of the growth media at 7.5 for both strains and under both growth conditions for at least 2 h past glucose

addition. Figure 1 Growth, glucose consumption and acetate build-up. Growth, glucose consumption and acetate formation in strain Newman (wt) and its isogenic ΔccpA mutant (ΔccpA). Cells were grown to an AZD1390 price OD600 of 1, cultures were split and 10 mM glucose was added to one half of the culture (squares), while the other half remained without glucose (triangles). Cells were sampled at an OD600 of 1 and 30 min after glucose addition for RNA isolation (indicated by arrows). Experiments shown are representative for three independent experiments.

Transcriptome analysis The total number of genes, which were expressed at a sufficient level to give meaningful data, was 2479. 111 of these genes had no homologues in strain Newman, and were therefore excluded from the analysis. Of the 2368 remaining Pregnenolone genes, a total of 155 were found to be affected upon glucose addition in a CcpA-dependent manner, while 21 genes seemed to be controlled by CcpA and other regulatory Selleck Vactosertib proteins at the same time in the presence of glucose, and 10 genes exhibited CcpA-independent glucose effects. The largest group, comprising 226 genes, however, was affected by ccpA inactivation even without glucose addition to the LB medium (Table 1). While regulatory classes partly overlapped, the overall range of differential gene expression was only narrow, peaking around 2- to 3-fold induction or repression. Table 1 Numbers of S.