Molecular cloning and functional analysis of the cysG and nirB genes of Escherichia coli K12, two closely-linked genes required for NADH-dependent nitrite reductase activity
✍ Scribed by Macdonald, Heather ;Cole, Jeff
- Publisher
- Springer
- Year
- 1985
- Tongue
- English
- Weight
- 703 KB
- Volume
- 200
- Category
- Article
- ISSN
- 0026-8925
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✦ Synopsis
We have cloned two genes, nirB + and eysG + which are required for NADH-dependent nitrite reductase to be active, from the 74 rain region of the Escherichia coli chromosome. Restriction mapping and complementation analysis establish the gene order crp-nirB-cysG-aroB. Both genes are trans-dominant in merodiploids and, under some conditions, can be expressed independently. The cysG + gene can be expressed from both high and low copy number plasmids carrying a 3.6 kb PstI-EcoRI restriction fragment. Attempts to sub-clone the nirB + gene into pBR322 on a 14.5 kb EeoRI fragment were unsuccessful, but this fragment was readily sub-cloned into and expressed from the low copy number plasmid pLG338 (Stoker et al. 1982). Overproduction of the 88 kDa nitrite reductase apoprotein by strains carrying a functional nirB + gene suggests that nirB is the structural gene for this enzyme. AB2847 BHB2688 BHB2690 DB6659 JCB213 JCB303 JCB312 JCB322 JCB323 JCB421 JCB422 JCB423 JCB424 JCB425 JCB426 JCB427 JCB429 JCB432 to 439 JCB512 RVChll M182 M182Acrp39 ar o B3 51malA N 20 5re c A ( 2 imm4 34cI tsb 2r ed3 EamSam 7) N205recA(2imm434cItsb2red3Daml 5Sam7) recAsrl: : TnlO cysG213 RVChlt nirB: : Mud I (ApRlac +) AB2847Mal + aroBcysG72 JCB303AnirB JCB303A (nirB-cysG)