title: Molecular dissection of the silkworm ribosomal stalk complex: the role of multiple copies of the stalk proteins.
abstract: In animal ribosomes, two stalk proteins P1 and P2 form a heterodimer, and the two dimers, with the anchor protein P0, constitute a pentameric complex crucial for recruitment of translational GTPase factors to the ribosome. To investigate the functional contribution of each copy of the stalk proteins, we constructed P0 mutants, in which one of the two C-terminal helices, namely helix I (N-terminal side) or helix II (C-terminal side) were unable to bind the P1-P2 dimer. We also constructed 'one-C-terminal domain (CTD) stalk dimers', P1-P2ΔC and P1ΔC-P2, composed of intact P1/P2 monomer and a CTD-truncated partner. Through combinations of P0 and P1-P2 variants, various complexes were reconstituted and their function tested in eEF-2-dependent GTPase and eEF-1α/eEF-2-dependent polyphenylalanine synthesis assays in vitro. Double/single-CTD dimers bound to helix I showed higher activity than that bound to helix II. Despite low polypeptide synthetic activity by a single one-CTD dimer, its binding to both helices considerably increased activity, suggesting that two stalk dimers cooperate, particularly in polypeptide synthesis. This promotion of activity by two stalk dimers was lost upon mutation of the conserved YPT sequence connecting the two helices of P0, suggesting a role for this sequence in cooperativity of two stalk dimers.
authors: Department of Biology, Faculty of Science, Niigata University, Nishi-ku, Ikarashi-2, Niigata 950-2181, Japan.
authors: Kentaro
authors: K
authors: Baba
authors: Kazuhiro
authors: K
authors: Tumuraya
authors: Isao
authors: I
authors: Tanaka
authors: Min
authors: M
authors: Yao
authors: Toshio
authors: T
authors: Uchiumi
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D012831:Bombyx; D017124:Conserved Sequence; D019476:Insect Proteins / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D055503:Protein Multimerization; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D019476:Insect Proteins; D012269:Ribosomal Proteins
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