Cloning and expression of the gene encoding the soluble cytochrome b562 of Escherichia coli.
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Nikkila H, Gennis RB, Sligar SG
Cloning and expression of the gene encoding the soluble cytochrome b562 of Escherichia coli.
Eur J Biochem. 1991 Dec 5;202(2):309-13.
- PubMed ID
- 1761034 [ View in PubMed]
- Abstract
The gene for the soluble cytochrome b562 from Escherichia coli B has been cloned on a SalI fragment. The analysis of the gene reveals the presence of a leader sequence in front of the sequence encoding the mature protein. Expression of cytochrome b562 using the lac-promoter produced the protein to a level of 3-5% of total protein. This over-production enables employment of a simple, high-yield purification protocol to obtain homogeneous cytochrome b562. Spectroscopic and N-terminal sequence analyses of the purified protein demonstrate that it is identical to the chromosomally expressed cytochrome b562 purified and characterized from E. coli B [Itagaki, E. & Hager, L.P. (1966) J. Biol. Chem. 241, 3687-3695]. It is demonstrated that the genomic sequence codes for a classic N-terminal signal sequence and that mature cytochrome b562 is translocated to the periplasmic space.