Abstract
Interference factors are characterized by their mRNA-specific effects on IF3 activity, in initiating the translation of various mRNA's. The assay purification and properties of interference factor i 2.3 (i-a) are described in this chapter. The assay is based on the fact that this protein inhibits translation of native MS2 RNA, but not T4 mRNA. The inhibition of MS2 RNA translation is because of the fact that factor i blocks ribosome binding and initiator fMet-tRNA binding at the coat protein initiation site, which is the only site exposed on native MS2 RNA. When the MS2 RNA is first unfolded by mild formaldehyde treatment, initiation can take place at the three cistrons independently, and because factor i blocks only coat protein synthesis, this template is actively translated in the presence of factor i. The assay of factor i is carried out by measuring the differential effect on at least two different mRNA templates.
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