Photochemical cross-linking of initiation factor-3 to Escherichia coli 30 S ribosomal subunits.

Initiation factor-3 has been photochemically cross-linked to Escherichia coli 30 S ribosomal subunits by means of near-ultraviolet (> 285 nm) irradiation. The cross-linking was judged to be specific since noncovalent binding was required for subsequent cross-linking and was prevented by the presence of 0.5 M NH4Cl or 0.5 mM aurintricarboxylic acid. Covalent attachment reached its maximum level of 8.5 to 11% of the noncovalently bound IF-3 after 60 min of irradation. Cross-linking was unaffected by the presence of the photosensitizer, acetone (5 or 10%), but was reduced to 30 to 40% of its maximum level by addition of 5 mM dithiothreitol, a free radical scavenger. Analysis of the 14C-labeled IF-3 x 30 S subunit covalent complex revealed that the IF-3 was distributed between the protein and RNA of the subunits in a 3:1 ratio. The target proteins have been identified as S7, S11, S12, S18, and S21 by immunochemical techniques.