Structure of Ribosomal Protein L6 from Escherichia coli

Abstract

Protein L6 from the 50 S ribosomal subunit of E. coli was investigated using fluorimetric techniques. The intrinsic fluorophore Trp-61 and fluorescent labels (acetylaminoethyl-dansyl and acetylaminofluorescein) attached to the residue Cys-124 were used. It proved possible to incorporate fluorescence-labeled L6 into the 50 S ribosome. Trp-61 is exposed to solvent, as shown by its emission wavelength and by quenching experiments; the latter also show that it lies in a pocket with a high positive charge due to the basic residues in the N-terminal fragment. Cys-124 lies in a less strongly positive region. Upon incorporation into the 50 S subunit, the label on Cys-124 becomes less accessible for quenching but its positive potential rises, showing the absence of direct contact with 23 S RNA. Analysis of anisotropy data indicates a considerable degree of asphericity of free L6. Energy transfer between Trp-61 and the dansyl label on Cys-124, measured by donor quenching and acceptor enhancement, reveals a separation of 3.5 .+-. 0.4 nm (35 .+-. 4 .ANG.) between fluorophores.