RsfA (YbeB) Proteins Are Conserved Ribosomal Silencing Factors
(A) Oligo(Phe) synthesis in a pure system containing pre-charged Phe-tRNAs (ten times over ribosomes), 30S and 50S subunits and the purified factors EF-Tu, EF-Ts and EF-G plus/minus RsfA from E. coli, 100% corresponds to 7 Phe incorporated per ribosome. Left panel, when indicated RsfA was added to the 50S subunits, before 30S subunits were added starting oligo(Phe) synthesis. Right panel, AcPhe-tRNA was bound to 70S ribosomes in the presence of poly(U) before the addition of RsfA. (B) Sister-aliquots from the same samples shown in (A) were analyzed on a sucrose gradient before oligo(Phe) synthesis. The presence of RsfA significantly reduces the fraction of 70S ribosomes. (C) Oligo(Phe)-synthesis as in (A) but with purified mitochondrial components (pig liver) and human mtRsfA (C7orf30). 39S and 28S indicate the large and small ribosomal subunits, 55S the associated mitochondrial ribosomes. For details see Experimental Procedures.