Escherichia coli strain MG1655 was chosen for sequencing because the few mutations it carries (ilvG rfb-50 rph-1) were considered innocuous. However, it has a number of growth defects. Internal pyrimidine starvation due to polarity of the rph-1 allele on pyrE was problematic in continuous culture. Moreover, the isolate of MG1655 obtained from the E. coli Genetic Stock Center also carries a large deletion around the fnr (fumaratenitrate respiration) regulatory gene. Although studies on DNA microarrays revealed apparent cross-regulation of gene expression between galactose and lactose metabolism in the Stock Center isolate of MG1655, this was due to the occurrence of mutations that increased lacY expression and suppressed slow growth on galactose. The explanation for apparent cross-regulation between galactose and N-acetylglucosamine metabolism was similar. By contrast, cross-regulation between lactose and maltose metabolism appeared to be due to generation of internal maltosaccharides in lactose-grown cells and may be physiologically significant. Lactose is of restricted distribution: it is normally found together with maltosaccharides, which are starch degradation products, in the mammalian intestine. Strains designated MG1655 and obtained from other sources differed from the Stock Center isolate and each other in several respects. We confirmed that use of other E. coli strains with MG1655-based DNA microarrays works well, and hence these arrays can be used to study any strain of interest. The responses to nitrogen limitation of two urinary tract isolates and an intestinal commensal strain isolated recently from humans were remarkably similar to those of MG1655.

The nag regulon located at 15.5 min on the Escherichia coli chromosome consists of two divergent operons, nagE and nagBACD, encoding genes involved in the uptake and metabolism of N-acetylglucosamine. Null mutations have been created in each of the genes by insertion of antibiotic resistance cartridges. The phenotypes of the strains carrying the insertions in nagE, B and A were consistent with the previous identification of gene products: nagE, EII(Nag), the N-acetylglucosamine specific transporter of the phosphotransferase system and nagB and nagA, the two enzymes necessary for the degradation of N-acetylglucosamine. Insertions in the nagC result in derepression of the nag genes, which is consistent with earlier observations that the nagC gene encodes the repressor of the regulon. Insertions in nagA also provoke a derepression, implying that nagA has a role in the regulation of the expression of the nag regulon as well as in the degradation of the amino-sugars. N-acetylglucosamine-6-phosphate, the intracellular product of N-acetylglucosamine transport and the substrate of the nagA gene product, is shown to be an inducer of the regulon and this suggests how nagA mutations result in derepression: the absence of N-acetylglucosamine-6-phosphate deacetylase allows N-acetylglucosamine-6-phosphate to accumulate and induce the regulon.

SummaryThe gene for the glucose-specific transporter of the phosphotransferase system, ptsG, is expressed from two promoters separated by 141 bp. The expression of the major, shorter transcript is very strongly dependent upon cAMP/CAP. However, unlike other CAPactivated genes, the expression of ptsG is higher in glucose media than in glycerol, implying that ptsG is controlled by a glucose-inducible regulator. A mutation in the mlc gene greatly enhances ptsG expression in a glycerol-grown culture but has no effect on ptsG expression during growth on glucose. The mlc gene encodes a transcriptional regulator that has been shown to affect the expression of manXYZ and malT. ptsG mRNA levels are lower in the mlc strain grown on glucose than in the same strain grown on glycerol. This is presumably because of the greater catabolite repression in the glucose culture than in glycerol. The final level of expression of ptsG in a mlc þ strain in glucose is a compromise between specific induction by glucose and generalized catabolite repression. The result is that ptsG expression is very similar in glucose-grown cultures of wild-type and mlc strains. The Mlc protein binds to two sites centred at ¹6 and ¹175 upstream of the major ptsG transcript. CAP binds at ¹40.5 compared with this site, typical of class II CAP-regulated promoters, and the binding of CAP and Mlc is co-operative.