title: Protein-RNA affinity of ribosomal protein L1 mutants does not correlate with the number of intermolecular interactions.
abstract: Ribosomal protein L1, as part of the L1 stalk of the 50S ribosomal subunit, is implicated in directing tRNA movement through the ribosome during translocation. High-resolution crystal structures of four mutants (T217V, T217A, M218L and G219V) of the ribosomal protein L1 from Thermus thermophilus (TthL1) in complex with a specific 80 nt fragment of 23S rRNA and the structures of two of these mutants (T217V and G219V) in the RNA-unbound form are reported in this work. All mutations are located in the highly conserved triad Thr-Met-Gly, which is responsible for about 17% of all protein-RNA hydrogen bonds and 50% of solvent-inaccessible intermolecular hydrogen bonds. In the mutated proteins without bound RNA the RNA-binding regions show substantial conformational changes. On the other hand, in the complexes with RNA the structures of the RNA-binding surfaces in all studied mutants are very similar to the structure of the wild-type protein in complex with RNA. This shows that formation of the RNA complexes restores the distorted surfaces of the mutant proteins to a conformation characteristic of the wild-type protein complex. Domain I of the mutated TthL1 and helix 77 of 23S rRNA form a rigid body identical to that found in the complex of wild-type TthL1 with RNA, suggesting that the observed relative orientation is conserved and is probably important for ribosome function. Analysis of the complex structures and the kinetic data show that the number of intermolecular contacts and hydrogen bonds in the RNA-protein contact area does not correlate with the affinity of the protein for RNA and cannot be used as a measure of affinity.
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Svetlana
authors: S
authors: Tishchenko
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Olga
authors: O
authors: Kostareva
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Azat
authors: A
authors: Gabdulkhakov
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Alisa
authors: A
authors: Mikhaylina
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Ekaterina
authors: E
authors: Nikonova
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Natalia
authors: N
authors: Nevskaya
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Alena
authors: A
authors: Sarskikh
authors: Biocenter, Division of Medical Biochemistry, Innsbruck Medical University, Innrain 80-82, 6020 Innsbruck, Austria.
authors: Wolfgang
authors: W
authors: Piendl
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Maria
authors: M
authors: Garber
authors: Institute of Protein Research, Russian Academy of Sciences, Institutskaya 4, 142290 Pushchino, Moscow Region, Russian Federation.
authors: Stanislav
authors: S
authors: Nikonov
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D017354:Point Mutation; D011487:Protein Conformation; D012338:RNA, Ribosomal, 23S / Q000737:chemistry / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016963:Thermus thermophilus / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D012338:RNA, Ribosomal, 23S; D012269:Ribosomal Proteins; C028546:ribosomal protein L1
keywords: kinetic experiments; protein–RNA interactions; ribosomal protein L1 mutants