PMID:8932712
| Citation |
Bradbury, AJ, Gruer, MJ, Rudd, KE and Guest, JR (1996) The second aconitase (AcnB) of Escherichia coli. Microbiology (Reading, Engl.) 142 ( Pt 2):389-400 |
|---|---|
| Abstract |
The second aconitase (AcnB) of Escherichia coli was partially purified from an acnA::kanR 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 center dot 85 min (131 center dot 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 lambdaacnB phages from the Kohara lambda-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 > or = 98% homogeneity from a genetically enriched source (JRG3171) and shown to be a monomeric protein of Mr 100 000 (SDS-PAGE) and 105 000 (gel filtration analysis) compared with Mr 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). |
| Links | |
| Keywords |
Aconitate Hydratase/chemistry; Aconitate Hydratase/genetics; Aconitate Hydratase/isolation & purification; Amino Acid Sequence; Bacteriophage lambda/genetics; Base Sequence; DNA Primers/genetics; DNA, Bacterial/genetics; Escherichia coli/enzymology; Escherichia coli/genetics; Gene Amplification; Genes, Bacterial; Molecular Sequence Data; Molecular Structure; Restriction Mapping; Sequence Homology, Amino Acid |
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