Editor: Roger Buxton
Use of a codon alteration strategy in a novel approach to cloning the Mycobacterium tuberculosis diaminopimelic acid epimerase
Article first published online: 4 JUL 2006
FEMS Microbiology Letters
Volume 262, Issue 1, pages 39–47, September 2006
How to Cite
Usha, V., Dover, L. G., Roper, D. L., Lloyd, A. J. and Besra, G. S. (2006), Use of a codon alteration strategy in a novel approach to cloning the Mycobacterium tuberculosis diaminopimelic acid epimerase. FEMS Microbiology Letters, 262: 39–47. doi: 10.1111/j.1574-6968.2006.00356.x
- Issue published online: 4 JUL 2006
- Article first published online: 4 JUL 2006
- Received 30 May 2006; revised 8 June 2006; accepted 13 June 2006.First published online 4 July 2006.
- diaminopimelic acid;
Previous attempts to express the diaminopimelate epimerase gene dapF of Mycobacterium tuberculosis in Escherichia coli resulted in undetectable enzyme yields. We used silent mutation of the first 10 codons of the recombinant ORF in an attempt to reduce the formation of secondary structures that might occur near the 5′ end of the mRNA and inhibit translation. This significantly increased the yield of the enzyme, which was purified and characterized biochemically. This strategy could be generally applied to other mycobacterial genes that are difficult to express hetero-specifically and here provided pure M. tuberculosis DapF, a good foundation for future research in antimycobacterial agents.