The 24,000-Dalton Cap-Binding Protein

The initial identification of a cap-binding protein resulted from experiments by Sonenberg and Shatkin (1977), who developed a crosslinking assay to detect polypeptides which were in physical proximity to the 5 ends of capped mRNAs. Reovirus mRNA was synthesized containing H-label in the methyl groups of the 5 cap. The mRNA was oxidized with NaI04 so as to convert the 2, 3 -cw-diol of the 5 -terminal m G to a reactive dialdehyde, and the oxidized mRNA was then incubated with ribosomal salt wash from rabbit reticulocytes or from Ehrlich ascites cells (Sonenberg et al., 1978). The 

The conditions of the crosslinking assay described above were modified to examine the effect of ATP and Mg on the crosslinking of reticulocyte initiation factors to mRNA caps (Sonenberg, 1981 Sonenberg et al., 1981). In addition to the 24-CBP, several other polypeptides having molecular weights of 28,000, 50,000, and 80,000 daltons were shown to specifically crosslink to the oxidized terminus of reovirus mRNA. Specific crosslinking of the latter polypeptides was totally dependent upon ATP and Mg , in contrast to crosslink- 

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