IDENTIFICATION OF A REGION OF BACILLUS-SUBTILIS FFH, A HOMOLOG OF MAMMALIAN SRP54 PROTEIN, THAT IS ESSENTIAL FOR BINDING TO SMALL CYTOPLASMIC RNA

Bacillus subtilis Ffh and scRNA are homologues of mammalian SRP54 and SR9 RNA, respectively, which are components of the eukaryotic signal r ecognition particle (SR9). Ffh (446 amino acids) interacts with scRNA to form a stable complex in vivo. Here, we identified an RNA-binding d omain of Ffh. The results obtained using a series of deletion mutants show that amino acid positions 364 to 432 in the C-terminal region of Ffh correlates with its ability to bind RNA. The amino acid sequence o f this region is well conserved among members of the SRP54 protein fam ily. This sequence contains two hydrophobic regions (h2, 364 to 391, a nd h3, 416 to 435), separated by the positively charged amino acid mot if, (398)RRKRIAKGSG(407). Among the basic amino acid residues in this region, Arg-401 was essential for binding to scRNA, but Arg-399 and Ly s-400 were not. The co-existence of Arg-398 and Lye-404 was necessary for the same affinity as wild type Ffh. The two glycine residues of th e (405)GSG(407) were also essential. MH23 peptide (91 amino acids) enc ompassing from 356 to 446, consisting of h2-RRKRIAKGSG-h3, bound scRNA with the same affinity as wild type Ffh, whereas a 24-amino acid synt hetic peptide (392)DIINASRRKRIAKGSGTSVQEVNR(415) did not. The region c ontaining two hydrophobic segments separated by the positively charged motif is the minimal requirement of Ffh for RNA binding.