Site-directed hydroxyl radical probing of the rRNA neighborhood of ribosomal protein S5.
Cysteine residues were introduced into three different positions distributed on the surface of ribosomal protein S5, to serve as targets for derivatization with an Fe(II)-ethyl-enediaminetetraacetic acid linker. Hydroxyl radicals generated locally from the tethered Fe(II) in intermediate ribonucleoprotein particles or in 30S ribosomal subunits reconstituted from derivatized S5 caused cleavage of the RNA, resulting in characteristically different cleavage patterns for the three different tethering positions. These findings provide constraints for the three-dimensional folding of 16S ribosomal RNA (rRNA) and for the orientation of S5 in the 30S subunit, and they further suggest that antibiotic resistance and accuracy mutations in S5 may involve perturbation of 16S rRNA.
Heilek
G M
GM
Center for Molecular Biology of RNA, Sinsheimer Laboratories, University of California, Santa Cruz 95064, USA.
Noller
H F
HF
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D003001:Cloning, Molecular; D003545:Cysteine / Q000737:chemistry; D004492:Edetic Acid / Q000031:analogs & derivatives; D004926:Escherichia coli; D005296:Ferrous Compounds / Q000737:chemistry; D017665:Hydroxyl Radical / Q000737:chemistry*; D008958:Models, Molecular; D015335:Molecular Probes; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D009942:Organometallic Compounds; D011487:Protein Conformation; D012335:RNA, Ribosomal / Q000737:chemistry*; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000187:drug effects; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000235:genetics; D000198:Spectinomycin / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D005296:Ferrous Compounds; D015335:Molecular Probes; D009942:Organometallic Compounds; D012335:RNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D012269:Ribosomal Proteins; C040469:ribosomal protein S30; C030831:ribosomal protein S5; D017665:Hydroxyl Radical; C042007:1-(4-bromoacetamidobenzyl)EDTA; D000198:Spectinomycin; D004492:Edetic Acid; D003545:Cysteine