* To whom correspondence should be addressed.
Received September 4, 2013
The task of the present work was to answer the question: is the free 5′-end needed for effective translation of a model polyribonucleotide template – polyuridylic acid – in a bacterial (E. coli) cell-free system? For this purpose, the template activities of the original polyuridylic acid with its free 5′-end and the polyuridylic acid with blocked 5′-end were compared in the bacterial cell-free translation system. To block the 5′-end, the cytidylic oligodeoxyribonucleotide with fluorescein residue at its 5′-end and uridylic oligoribonucleotide sequence at its 3′-end, schematically described as FAM(dC)10(rU)50, was covalently attached (ligated) to the 5′-end of the template polyuridylic acid. It was shown that the efficiency of polyphenylalanine synthesis on the 5′-blocked template and on the polyuridylic acid with free 5′-end was virtually the same. It was concluded that bacterial ribosomes are capable of effectively initiating translation at the polyuridylic sequence independently of the 5′-end of template polyribonucleotide, i.e. via an internal initiation mechanism, in the absence of a Shine–Dalgarno sequence and AUG start codon.
KEY WORDS: polyuridylic acid, translation initiation, polyphenylalanine synthesis, T4 RNA ligase, cell-free translation