Pathways for the utilization of N-acetyl-galactosamine and galactosamine in Escherichia coli

Among enteric bacteria, the ability to grow on N-acetyl-galactosamine (GalNAc or Aga) and on D-galactosamine (GalN or Gam) differs. Thus, strains B, C and EC3132 of Escherichia coli are Aga+ Gam+ whereas E. coli K-12 is Aga- Gam-, similarly to Klebsiella pneumoniae KAY2026, Klebsiella oxytoca M5a1 and Salmonella typhimurium LT2. The former strains carry a complete aga/kba gene cluster at 70.5 min of their gene map. These genes encode an Aga-specific phosphotransferase system (PTS) or IIAga (agaVWE) and a GalN-specific PTS or IIGam (agaBCD). Both PTSs belong to the mannose-sorbose family, i.e. the IIB, IIC and IID domains are encoded by different genes, and they share a IIA domain (agaF). Furthermore, the genes encode an Aga6P-deacetylase (agaA), a GalN6P deaminase (agaI), a tagatose-bisphosphate aldolase comprising two different peptides (kbaYZ) and a putative isomerase (agaS), i.e. complete pathways for the transport and degradation of both amino sugars. The genes are organized in two adjacent operons (kbaZagaVWEFA and agaS kbaYagaBCDI) and controlled by a repressor AgaR. Its gene agaR is located upstream of kbaZ, and AgaR responds to GalNAc and GalN in the medium. All Aga- Gam- strains, however, carry a deletion covering genes agaW' EF 'A; consequently they lack active IIAga and IIGam PTSs, thus explaining their inability to grow on the two amino sugars. Remnants of a putative recombination site flank the deleted DNA in the various Aga- Gam- enteric bacteria. Derivatives with an Aga+ Gam- phenotype can be isolated from E. coli K-12. These retain the DeltaagaW' EF 'A deletion and carry suppressor mutations in the gat and nag genes for galactitol and N-acetyl-glucosamine metabolism, respectively, that allow growth on Aga but not on GalN.     

Purification and characterization of the DNA-binding protein DnrI, a transcriptional factor of daunorubicin biosynthesis in Streptomyces peucetius

The DnrI protein, essential for the biosynthesis of daunorubicin in Streptomyces peucetius , was purified almost to homogeneity from dnrI expression strains of Escherichia coli and S. peucetius through several steps of chromatography. The proteins purified from both organisms had identical chromatographic and electrophoretic behaviour. Purified His-tagged or native DnrI was used to conduct DNA-binding assays by gel mobility-shift analysis, and the results showed no significant difference in the DNA-binding activity of native or His-tagged proteins. DnrI binds specifically to DNA segments containing the intergenic regions separating the putative dnrG–dpsABCD and dpsEF operons, and the dnrC gene and dnrDKPSQ operon. DNase I footprinting assays indicated that the DNA-binding sites for DnrI extended from upstream of the −10 to −35 regions of the dnrG or dpsE promoters to include about 65 bp of the dnrG – dpsE intergenic region and about 80 bp of the dnrC – dnrD intergenic region. Both binding sites contain imperfect inverted repeat sequences of 6–10 bp with a 5'-TCGAG-3' consensus sequence that was present in 4 out of 10 other promoter regions in the cluster of daunorubicin biosynthesis genes.     

The primary structure of the DeoR repressor from Escherichia coli K-12.

The nucleotide sequence of the deoR gene of E. coli, which codes for the DeoR repressor, has been determined. This gene codes for a polypeptide that is 252 amino acids residues in length. Computer-assisted analysis of the nucleotide sequence strongly suggests that the DNA binding domain of the DeoR repressor is located in the N-terminal part of the protein. After the coding region there is a dyad symmetry similar to a palindromic unit present outside many structural genes on the E. coli chromosome.