Green P J, Inouye M
J Mol Biol. 1984 Jul 5;176(3):431-42. doi: 10.1016/0022-2836(84)90499-6.
OmpA protein, a major outer membrane protein of Escherichia coli, is synthesized from a messenger RNA containing a 134-nucleotide 5' leader region. The role of this leader region in efficient ompA expression was investigated using a series of ompA-lacZ fusion plasmids. These plasmids differ in the amount of DNA encoding the ompA leader region which is fused to the lacZ structural gene. The fusion containing all but six nucleotides of the ompA leader produced the highest beta-galactosidase activity, while the fusion containing the shortest leader synthesized only 4% as much beta-galactosidase. Fusions with leaders intermediate in length produced between 6% and 24% of the activity found in the most efficient fusion. Quantitation of lacZ mRNA synthesis by DNA-RNA hybridization revealed differences in lacZ mRNA production reflecting the observed differences in beta-galactosidase activity. The primary effect of the ompA leader in maintaining high levels of mRNA is discussed in terms of the roles of mRNA secondary structure.
外膜蛋白A(OmpA)是大肠杆菌的一种主要外膜蛋白,由一种含有134个核苷酸的5'前导区的信使核糖核酸(mRNA)合成。使用一系列ompA - 乳糖酶(lacZ)融合质粒研究了该前导区在高效ompA表达中的作用。这些质粒在与lacZ结构基因融合的编码ompA前导区的DNA量上有所不同。包含ompA前导区除六个核苷酸外所有序列的融合体产生了最高的β - 半乳糖苷酶活性,而包含最短前导区的融合体合成的β - 半乳糖苷酶仅为最高活性融合体的4%。长度介于两者之间的前导区融合体产生的活性为最有效融合体的6%至24%。通过DNA - RNA杂交对lacZ mRNA合成进行定量分析,结果显示lacZ mRNA产量存在差异,这反映了观察到的β - 半乳糖苷酶活性差异。从mRNA二级结构的作用方面讨论了ompA前导区在维持高水平mRNA中的主要作用。