Dynamic pathways of -1 translational frameshifting #MMPMID24919156
Chen J; Petrov A; Johansson M; Tsai A; O?Leary SE; Puglisi JD
Nature 2014[Aug]; 512 (7514): 328-32 PMID24919156show ga
Spontaneous changes in the reading frame of translation are rare (frequency of 10?3 ? 10?4 per codon)1, but can be induced by specific features in the messenger RNA (mRNA). In the presence of mRNA secondary structures, a heptanucleotide ?slippery sequence? usually defined by the motif X XXY YYZ, and (in some prokaryotic cases) mRNA sequences that base pair with the 3? end of the 16S ribosomal rRNA (internal Shine-Dalgarno (SD) sequences), there is an increased probability that a specific programmed change of frame occurs, wherein the ribosome shifts one nucleotide backwards into an overlapping reading frame (?1 frame) and continues by translating a new sequence of amino acids2,3. Despite extensive biochemical and genetic studies, there is no clear mechanistic description for frameshifting. Here, we apply single-molecule fluorescence to track the compositional and conformational dynamics of the individual ribosomes at each codon during translation of a frameshift-inducing mRNA from the dnaX gene in Escherichia coli. Ribosomes that frameshift into the ?1 frame are characterized by a 10-fold longer pause in elongation compared to non-frameshifted ribosomes, which translate through unperturbed. During the pause, interactions of the ribosome with the mRNA stimulatory elements uncouple EF-G catalyzed translocation from normal ribosomal subunit reverse-rotation, leaving the ribosome in a non-canonical intersubunit rotated state with an exposed codon in the aminoacyl-tRNA site (A site). tRNALys sampling and accommodation to the empty A site either lead to the slippage of the tRNAs into the ?1 frame or maintain the ribosome into the 0 frame. Our results provide a general mechanistic and conformational framework for ?1 frameshifting, highlighting multiple kinetic branchpoints during elongation.
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Nature 2014 ; 512 (7514): 328-32
