Toeprint Assays for Detecting RNA Structure and Protein–RNA Interactions

Toeprint assays are primer extension inhibition assays that can detect the 3' end of an RNA secondary structure, the position of a bound RNA binding protein, as well as the position of a bound 30S ribosomal subunit or a stalled ribosome. Here we describe how this assay was used to identify an RNA hairpin that sequesters a Shine-Dalgarno sequence, how the RNA-binding protein CsrA can alter RNA structure and affect 30S ribosomal subunit binding, and how the macrolide antibiotic tylosin can induce ribosome stalling.