Effect of the relative position of the UGA codon to the unique secondary structure in the fdhF mRNA on its decoding by selenocysteinyl tRNA in Escherichia coli.

The fdhF mRNA for formate dehydrogenase H of Escherichia coli contains a UGA codon at position 140. This termination codon is decoded by selenocysteinyl tRNA (the selC product) with the aid of its own specific elongation factor, SelB. For this decoding, a unique secondary structure immediately downstream of the UGA codon has been shown to be essential (Zinoni, F., Heider, J., and Böck, A. (1990) Proc. Natl. Acad. Sci. U. S. A. 87, 4660-4664). We examined the positional effect of the UGA codon relative to the secondary structure on its decoding using a fdhF-lacZ fusion gene. When the UGA codon was separated by one codon (position -1) from the secondary structure, the UGA decoding, as measured by the beta-galactosidase activity, dropped to approximately 76% of the normal level but was still almost as fully dependent upon selC and selenium in the culture medium as in the case of the UGA codon in the normal position (position 0). However, when the UGA codon was separated by two codons (position -2), the decoding level further dropped to 20% of the normal level, and in addition, became dependent only on selC but independent of selenium. When the UGA codon was further separated by three codons (position -3), the decoding level of UGA (-3) became higher than the decoding of UGA (-2) and was completely independent from selC and selenium, indicating that the UGA codon was nonspecifically suppressed. A similar nonspecific suppression was observed for the UGA codon at position -4, but at a lower level. When two UGA codons were tandemly placed at positions 0 and -1, they were still able to be decoded at 17% of the normal level in a selC- and selenium-dependent manner. In the absence of the SelB function, the decoding level of UGA(0) dropped to 1.6% of the normal level, whereas the UGA(-1) decoding dropped to 7.5%. These results indicate that the UGA codon at position 0 is not only most effectively decoded by selenocysteinyl tRNA but also tightly blocked from its nonspecific suppression in the absence of any components required for the decoding.