Protein-rRNA binding features and their structural and functional implications in ribosomes as determined by cross-linking studies.
We have investigated protein-rRNA cross-links formed in 30S and 50S ribosomal subunits of Escherichia coli and Bacillus stearothermophilus at the molecular level using UV and 2-iminothiolane as cross-linking agents. We identified amino acids cross-linked to rRNA for 13 ribosomal proteins from these organisms, namely derived from S3, S4, S7, S14, S17, L2, L4, L6, L14, L27, L28, L29 and L36. Several other peptide stretches cross-linked to rRNA have been sequenced in which no direct cross-linked amino acid could be detected. The cross-linked amino acids are positioned within loop domains carrying RNA binding features such as conserved basic and aromatic residues. One of the cross-linked peptides in ribosomal protein S3 shows a common primary sequence motif--the KH motif--directly involved in interaction with rRNA, and the cross-linked amino acid in ribosomal protein L36 lies within the zinc finger-like motif of this protein. The cross-linked amino acids in ribosomal proteins S17 and L6 prove the proposed RNA interacting site derived from three-dimensional models. A comparison of our structural data with mutations in ribosomal proteins that lead to antibiotic resistance, and with those from protein-antibiotic cross-linking experiments, reveals functional implications for ribosomal proteins that interact with rRNA.
Urlaub
H
H
Max-Delbrück-Centrum für Molekulare Medizin, Berlin, Germany.
Kruft
V
V
Bischof
O
O
Müller
E C
EC
Wittmann-Liebold
B
B
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents; D001411:Geobacillus stearothermophilus / Q000737:chemistry*; D007096:Imidoesters; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011485:Protein Binding; D012336:RNA, Ribosomal, 16S / Q000737:chemistry* / Q000378:metabolism; D012338:RNA, Ribosomal, 23S / Q000737:chemistry* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry*; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D007096:Imidoesters; D010446:Peptide Fragments; D012336:RNA, Ribosomal, 16S; D012338:RNA, Ribosomal, 23S; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins; C005905:methyl 4-mercaptobutyrimidate