Publications

• Isolation and properties of N epsilon-hydroxylysine:acetyl coenzyme A N epsilon-transacetylase from Escherichia coli pABN11.

  Coy M., Paw B.H., Bindereif A., Neilands J.B.

  The enzyme N epsilon-hydroxylysine acetylase has been isolated from Escherichia coli 294 carrying recombinant plasmid ABN11. Activity of the enzyme was followed by measurement of the rate of appearance of 2-nitro-5-thiobenzoate, the product of cleavage of 5,5'-dithiobis(2-nitrobenzoate) by free co ... >> More

  The enzyme N epsilon-hydroxylysine acetylase has been isolated from Escherichia coli 294 carrying recombinant plasmid ABN11. Activity of the enzyme was followed by measurement of the rate of appearance of 2-nitro-5-thiobenzoate, the product of cleavage of 5,5'-dithiobis(2-nitrobenzoate) by free coenzyme A released from its acetyl derivative. The enzyme bound firmly to Reactive Blue 2-Sepharose CL-6B and was eluated with 1.5 M KCl. The protein gave a single band, corresponding to a Mr of 33,000, on polyacrylamide gel electrophoresis in sodium dodecyl sulfate. In contrast, gel filtration of the native enzyme gave a Mr of 150,000-200,000. A sequence analysis of the DNA at the junction of the first and second genes in the aerobactin operon, considered in conjunction with the N-terminal amino acid sequence of the isolated protein, enabled the conclusion that the acetylase is specified by the second gene in the complex. The enzyme transfers the acetyl moiety from acetyl coenzyme A to a variety of hydroxylamines, with N epsilon-hydroxylysine as the preferred substrate. In agreement with the results found by affinity chromatography, Coomassie Blue was observed to act as a potent inhibitor. << Less

  Biochemistry 25:2485-2489(1986) [PubMed] [EuropePMC]

• Aerobactin biosynthesis and transport genes of plasmid ColV-K30 in Escherichia coli K-12.

  de Lorenzo V., Bindereif A., Paw B.H., Neilands J.B.

  The iron-regulated aerobactin operon, about 8 kilobase pairs in size, of the Escherichia coli plasmid ColV-K30 was shown by deletion and subcloning analyses to consist of at least five genes for synthesis (iuc, iron uptake chelate) and transport (iut, iron uptake transport) of the siderophore. The ... >> More

  The iron-regulated aerobactin operon, about 8 kilobase pairs in size, of the Escherichia coli plasmid ColV-K30 was shown by deletion and subcloning analyses to consist of at least five genes for synthesis (iuc, iron uptake chelate) and transport (iut, iron uptake transport) of the siderophore. The gene order iucABCD iutA was established. The genes were mapped within restriction nuclease fragments of a cloned 16.3-kilobase-pair HindIII fragment. Stepwise deletion and subsequent minicell analysis of the resulting plasmids allowed assignment of four of the five genes to polypeptides of molecular masses 63,000, 33,000 53,000, and 74,000 daltons, respectively. The 74-kilodalton protein, the product of gene iutA, is the outer membrane receptor for ferric aerobactin, whereas the remaining three proteins are involved in biosynthesis of aerobactin. The 33-kilodalton protein, the product of gene iucB, was identified as N epsilon-hydroxylysine:acetyl coenzyme A N epsilon-transacetylase (acetylase) by comparison of enzyme activity in extracts from various deletion mutants. The 53-kilodalton protein, the product of gene iucD, is required for oxygenation of lysine. The 63-kilodalton protein, the product of gene iucA, is assigned to the first step of the aerobactin synthetase reaction. The product of gene iucC, so far unidentified, performs the second and final step in this reaction. This is based on the chemical characterization of two precursor hydroxamic acids (N epsilon-acetyl-N epsilon-hydroxylysine and N alpha-citryl-N epsilon-acetyl-N epsilon-hydroxylysine) isolated from a strain carrying a 0.3-kilobase-pair deletion in the iucC gene. The results support the existence of a biosynthetic pathway in which aerobactin arises by oxygenation of lysine, acetylation of the N epsilon-hydroxy function, and condensation of 2 mol of the resulting aminohydroxamic acid with citric acid. << Less

  J. Bacteriol. 165:570-578(1986) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.