Overproduction of the Bradyrhizobium japonicum c-type cytochrome subunits of the cbb3 oxidase in Escherichia coli.
We report on a system to improve expression of mature c-type cytochromes in Escherichia coli. It is based on the use of plasmid pEC86 that expresses the E. coli cytochrome c maturation genes ccmABCDEFGH constitutively, whereby the production of both endogenous and foreign c-type cytochromes was increased substantially. The periplasmic soluble domains of the c-type cytochrome subunits FixO and FixP of the Bradyrhizobium japonicum cbb3 oxidase could be expressed in E. coli only when pEC86 was provided in a degP-deficient strain. This shows that a stimulation of heme attachment by the Ccm maturase system combined with the diminished proteolytic activity in the periplasm can increase c-type cytochrome yields.
Arslan
E
E
Mikrobiologisches Institut, Eidgenössische Technische Hochschule, Schmelzbergstrasse 7, Zürich, CH-8092, Switzerland.
Schulz
H
H
Zufferey
R
R
Künzler
P
P
Thöny-Meyer
L
L
D020369:Bradyrhizobium / Q000201:enzymology*; D003574:Cytochrome c Group / Q000096:biosynthesis* / Q000235:genetics; D003576:Electron Transport Complex IV / Q000096:biosynthesis* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics; D010088:Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012995:Solubility
D003574:Cytochrome c Group; D008565:Membrane Proteins; D011994:Recombinant Proteins; C080743:cytochrome FixO; C080742:cytochrome FixP; D010088:Oxidoreductases; C020094:duroquinol oxidase; D003576:Electron Transport Complex IV