Self-initiation of translation of mRNAs devoid of translational initiatorsin Escherichia coli

Recent studies have shown that the canonical SD-anti-SD interaction is disp ensable for the initiation of translation of certain mRNAs in Escherichia c oli. In this study the cat and tetR genes were modified to either destroy c omplementarity to E. coli 16S RNA or completely delete their 5' non-transla ted regions. Thus a series of cat- and tetR-derived genes were constructed, cloned under a strong constitutive promoter and expressed in E. coli cells . The efficiency of expression was evaluated by the yield of CAT (for the c at gene) and cell viability in increasing concentrations of antibiotic (for both cat and tetR genes). The obtained results show that the mRNAs transcr ibed from both series of reporter genes (cat and tetR) were active in vivo. Their activity was preserved even in the cases when the length of their 5' non-translated leader sequences was reduced to one nucleotide for the cat gene and eight nucleotides for the tetR gene. The yield of protein obtained with the latter constructs was detectable and sufficient for bacteria to s urvive at 50-100 mug/ml chloramphenicol and 20 mug/ml tetracycline, respect ively.