The second aconitase (AcnB) of Escherichia coli

Alan J. Bradbury, Megan J. Gruer, Kenneth E. Rudd, John R. Guest

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The second aconitase (AcnB) of Escherichia coli was partially purified from an acnA::kan(R) mutant lacking AcnA, and the corresponding polypeptide identified by activity staining and weak cross-reactivity with AcnA antiserum. The acnB gene was located at 2.85 min (131.6 kb) in a region of the chromosome previously assigned to two unidentified ORFs. Aconitase specific activities were amplified up to fivefold by infection with λacnB phages from the Kohara λ-E. coli gene library, and up to 120-fold (50% of soluble protein) by inducing transformants containing a plasmid (pGS783) in which the acnB coding region is expressed from a regulated T7 promoter. The AcnB protein was purified to ≥ 98% homogeneity from a genetically enriched source (JRG3171) and shown to be a monomeric protein of M(r) 100,000 (SDS-PAGE) and 105,000 (gel filtration analysis) compared with M(r) 93,500 predicted from the nucleotide sequence. The sequence identity between AcnA and AcnB is only 17% and the domain organization of AcnA and related proteins (1-2-3-linker-4) is rearranged in AcnB (4-1-2-3).

Original languageEnglish
Pages (from-to)389-400
Number of pages12
JournalMicrobiology
Volume142
Issue number2
StatePublished - Feb 1 1996
Externally publishedYes

Fingerprint

Aconitate Hydratase
Escherichia coli
Proteins
Coliphages
Gene Library
Open Reading Frames
Gel Chromatography
Immune Sera
Polyacrylamide Gel Electrophoresis
Plasmids
Chromosomes
Staining and Labeling
Peptides
Infection
Genes

Keywords

  • Aconitase
  • Citric acid cycle
  • Escherichia coli
  • Iron-sulphur proteins
  • Protein domains

ASJC Scopus subject areas

  • Microbiology

Cite this

Bradbury, A. J., Gruer, M. J., Rudd, K. E., & Guest, J. R. (1996). The second aconitase (AcnB) of Escherichia coli. Microbiology, 142(2), 389-400.

The second aconitase (AcnB) of Escherichia coli. / Bradbury, Alan J.; Gruer, Megan J.; Rudd, Kenneth E.; Guest, John R.

In: Microbiology, Vol. 142, No. 2, 01.02.1996, p. 389-400.

Research output: Contribution to journalArticle

Bradbury, AJ, Gruer, MJ, Rudd, KE & Guest, JR 1996, 'The second aconitase (AcnB) of Escherichia coli', Microbiology, vol. 142, no. 2, pp. 389-400.
Bradbury AJ, Gruer MJ, Rudd KE, Guest JR. The second aconitase (AcnB) of Escherichia coli. Microbiology. 1996 Feb 1;142(2):389-400.
Bradbury, Alan J. ; Gruer, Megan J. ; Rudd, Kenneth E. ; Guest, John R. / The second aconitase (AcnB) of Escherichia coli. In: Microbiology. 1996 ; Vol. 142, No. 2. pp. 389-400.
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