Linkage map of Escherichia coli K-12, edition 8.

Linkage map of Escherichia coli K-12, edition 6.

Identifying gene targets for the metabolic engineering of lycopene biosynthesis in Escherichia coli.

The nucleotide sequence of a 1980-base-pair segment of DNA, containing the lpd gene encoding the lipoamide dehydrogenase component (E3) of the pyruvate dehydrogenase complex of Escherichia coli K 12, has been determined by the dideoxy chain-termination method. The lpd structural gene comprises 1419 base pairs (473 codons, excluding the initiating AUG codon). It is preceded by a good promoter and an excellent ribosome binding site and it ends with a typical rho-independent terminator sequence. The results confirm that the lpd gene is an independent gene linked to, but not part of, the ace operon that encodes the E1 and E2 components of the pyruvate dehydrogenase complex. The location and transcriptional polarity of the lpd gene relative to the restriction map of the corresponding region of DNA, are completely consistent with previous genetic and post-infection labelling studies. The composition, Mr (50554 or 51274 if the FAD cofactor is included), aminoterminal sequence and carboxy-terminal sequence predicted from the nucleotide sequence are in excellent agreement with previous studies on the purified enzyme. The enzyme also exhibits a remarkable degree of sequence homology with peptides of the pig heart enzyme and with other pyridine nucleotide disulphide oxidoreductases whose sequences have been defined: human erythrocyte glutathione reductase and plasmid-encoded mercuric reductase.

Nucleotide sequence of the lipoamide dehydrogenase gene of Escherichia coli K12.

This map is an update of the edition 9 map by Berlyn et al. (M. K. B. Berlyn, K. B. Low, and K. E. Rudd, p. 1715–1902, in F. C. Neidhardt et al., ed., Escherichia coli and Salmonella: cellular and molecular biology, 2nd ed., vol. 2, 1996). It uses coordinates established by the completed sequence, expressed as 100 minutes for the entire circular map, and adds new genes discovered and established since 1996 and eliminates those shown to correspond to other known genes. The latter are included as synonyms. An alphabetical list of genes showing map location, synonyms, the protein or RNA product of the gene, phenotypes of mutants, and reference citations is provided. In addition to genes known to correspond to gene sequences, other genes, often older, that are described by phenotype and older mapping techniques and that have not been correlated with sequences are included.

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Linkage Map of Escherichia coli K-12, Edition 10: The Traditional Map

Twenty-eight spontaneous auxotrophic aroP mutants with deletions in the azi-nadC-aroP-aceE-aceF-lpd region of the Escherichia coli K 12 chromosome were characterized genetically with respect to various azi, nadC, ace and lpd markers by P1-mediated transduction. One mutant (K δ18; aroP-lpdδ) had a deletion which extended through the aceE and aceF genes to end within the lpd gene. The polarity of the ace operon (aceE to aceF) was confirmed. It was concluded that 10 out of 15 deletions generating a strict requirement for acetate terminated in the aceE gene. Of the ten, three mutants (K δ22, cδ41 and cδ42) synthesized detectable dihydrolipoamide acetyltransferase (the aceF gene product) and seven were assumed to possess deletions generating polar effects on aceF gene expression. Five deletions appeared to extend into the aceF gene. A further five deletions, which limited the expression of the ace operon without generating an Ace- phenotype or a complete Ace- phenotype, ended closest to the aroP-proximal aceE markers. The opposite ends of all these deletions appeared to terminate before (10), within (2) or extend beyond (9) the nadC gene. There was no obvious correlation between the deletion end-points and the corresponding lipoamide dehydrogenase activities, which ranged from 30 to 95% of parental levels in different deletion strains. The remaining seven deletions simply extended between the aroP and nadC genes (nad-aroPδ) without affecting expression of the ace operon.
Regulation of the synthesis of the pyruvate and α-ketoglutarate dehydrogenase complexes was investigated in some of the parental and deletion strains under different physiological conditions including thiamin-deprivation. The results indicate that the syntheses of the two dehydrogenase complexes are independently regulated. Expression of the lpd gene appears to be coupled to complex synthesis but can be dissociated under some conditions. Mechanisms for regulating lpd gene expression are discussed and an autogenous mechanism involving uncomplexed lipoamide dehydrogenase functioning as a negatively acting repressor at the operator site of an independent lpd gene is proposed as the simplest mechanism which is consistent with all available information.

Biochemical genetics of the alpha-keto acid dehydrogenase complexes of Escherichia coli K12: genetic characterization and regulatory properties of deletion mutants.

D. Langley

Papers

Biochemical genetics of the alpha-keto acid dehydrogenase complexes of Escherichia coli K12: genetic characterization and regulatory properties of deletion mutants.