%0 Journal Article %A Bradbury, Alan J. %A Gruer, Megan J. %A Rudd, Kenneth E. %A Guest, John R. %T The second aconitase (AcnB) of Escherichia coli %D 1996 %J Microbiology, %V 142 %N 2 %P 389-400 %@ 1465-2080 %R https://doi.org/10.1099/13500872-142-2-389 %K aconitase %K protein domains %K Escherichia coli %K iron-sulphur proteins %K citric acid cycle %I Microbiology Society, %X Summary: 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 100000 (SDS-PAGE) and 105000 (gel filtration analysis) compared with M r 93500 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). %U https://www.microbiologyresearch.org/content/journal/micro/10.1099/13500872-142-2-389