Over-expression of Escherichia coli F1Fo–ATPase subunit a is inhibited by instability of the uncB gene transcript
详细信息    查看全文
文摘
Little is known about the stability of transcripts encoding membrane proteins in strong expression systems and its effect on membrane protein over-production. We have expressed all the genes encoding subunits of the membrane domain Fo of the ATP synthase in a T7 RNA polymerase-based system. All of them but uncB (subunit a) were expressed separately at very high levels in the bacterial hosts Escherichia coli C41(DE3) and C43(DE3). However, expression of uncB was extremely toxic to the bacteria. Northern blot analysis showed that the level of accumulation of the mRNA from uncB was very low. Deletion of uncB in combination with gene fusion experiments demonstrated that the middle region of the gene, encoding amino acids 92–171, exhibited a dominant toxic phenotype associated with a very poor level of expression. Green fluorescent protein fusions with N- and C-ends of uncB helped to stabilize the mRNA and to obtain high yields of protein.

© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号

地址:北京市海淀区学院路29号 邮编:100083

电话:办公室:(+86 10)66554848;文献借阅、咨询服务、科技查新:66554700