Efficient removal of sulfide following integration of multiple copies of the sulfide-quinone oxidoreductase gene (sqr) into the Escherichia coli chromosome
H. Shibata et al., Efficient removal of sulfide following integration of multiple copies of the sulfide-quinone oxidoreductase gene (sqr) into the Escherichia coli chromosome, J BIOSCI BI, 91(5), 2001, pp. 493-499
For the oxidation and removal of hydrogen sulfide, which causes an offensiv
e odor from the contents of animal intestines, recombinant strains of Esche
richia coli were constructed. The sulfide-quinone oxidoreductase gene (sqr)
from Rhodobacter capsulatus was integrated in low copy numbers into the ch
romosome of Escherichia coli W3110. Multiple copies of sqr on plasmids were
also delivered into the cytoplasm of the same strain. The sqr genes were h
omologously transducted onto the chromosomal lacZ region and their existenc
e there was verified by Southern blot analysis. Sulfide oxidation in a chem
ical medium effectively increased for the recombinant strains which carried
2 similar to3 copies of sqr under the control of the lac or tac promoter i
n the chromosome, and also for strains which carried 10 copies of sqr under
the control of the lac or tac promoter on plasmids. In both types of recom
binant, the tac promoter was more effective for SQR expression than the lac
promoter. Construction of a recombinant with 3 copies of sqr under the con
trol of the tac promoter in the chromosome was unsuccessful. In recombinant
s with SQR activity lower than 700 nmol/mg cell protein/min, oxygen consump
tion increased proportionally to SQR activity. An elevation in SQR activity
in this range resulted in an increase in oxygen consumption and a decrease
in sulfide concentration. When the recombinant cells were cultured until t
he 160th generation, WL2, WL3 and WT2, which carried 2, 3 and 2 copies of s
qr in the chromosome, respectively, retained SQR activity similar to that o
f the first generation. For WL300 and WT20 which carried multi-copies of sq
r in plasmids SQR activity was undetectable. The recombinant with 2 copies
of sqr in the chromosome regulated by the tac promoter was most suitable fo
r sulfide oxidation and growth of the cells.