ABSTRACT
In this work we show for the first time that the overproduced N-terminal fragment (residues 1-91) of ribosomal protein TL5 binds specifically to 5S rRNA and that the region of this fragment containing residues 80-91 is a necessity for its RNA-binding activity. The fragment of Escherichia coli 5S rRNA protected by TL5 against RNase A hydrolysis was isolated and sequenced. This 39 nucleotides fragment contains loop E and helices IV and V of 5S rRNA. The isolated RNA fragment forms stable complexes with TL5 and its N-terminal domain. Crystals of TL5 in complex with the RNA fragment diffracting to 2.75 A resolution were obtained.
Subject(s)
Bacterial Proteins/metabolism , RNA, Ribosomal, 5S/metabolism , RNA-Binding Proteins/metabolism , Ribosomal Proteins/metabolism , Thermus thermophilus/metabolism , Amino Acid Sequence , Binding Sites , Molecular Sequence Data , Nucleic Acid Conformation , RNA, Ribosomal, 5S/chemistry , Sequence Homology, Amino AcidABSTRACT
The gene encoding the 5S rRNA-binding ribosomal protein TL5 from Thermus thermophilus, an extremely thermophilic species, was expressed in E. coli. A method for isolation of TL5 from the overproducing strain was developed. Samples of TL5 protein isolated from ribosomes and the overproducing strain displayed identical RNA-binding properties. Circular dichroic spectroscopy was used to calculate the secondary structure of the protein. TL5 was shown to form a stable complex with the 3'-terminal fragment of 5S rRNA, which is similar to the fragment of E. coli RNA that binds to L25 protein. The data suggest that TL5 from T. thermophilus and L25 from E. coli bind to similar sites on the 5S rRNA molecule.
