In bacteria and yeast, programmed frameshift signals can have rather different elements, as, e.g. the upstream Shine-Dalgarno binding element in the autoregulatory RF2 gene frameshift site first described in Escherichia coli (11) or the different pattern of the +1 frameshift stimulating heptanucleotide sequences present in Saccharomyces Ty elements (2) . However, many frameshift signals deviate little from those described for the virus-derived system used here and many signals are of such general character that ribosomes from different kingdoms of life will respond to them by shifting frame (12) . This happens not always with the same efficiency as in the original organism (12, 13) and there are even examples found where a frameshift element can direct the ribosomes into À2 or +1 frameshift depending on the test organism (14) . Here, we challenged E. coli ribosomes by constructing artificial frameshifting signals containing pseudoknot-like structures with strong stems. Using a refined frameshift assay, involving two-dimensional (2D) gel electrophoresis of pulse labelled proteins, we show that a significant amount of frameshifted ribosomes permanently stall within the strongest pseudoknots which therefore efficiently act as roadblocks.