Interaction among silkworm ribosomal proteins P1, P2 and P0 required for functional protein binding to the GTPase-associated domain of 28S rRNA.
Acidic ribosomal phosphoproteins P0, P1 and P2 were isolated in soluble form from silkworm ribosomes and tested for their interactions with each other and with RNA fragments corresponding to the GTPase-associated domain of residues 1030-1127 (Escherichia coli numbering) in silkworm 28S rRNA in vitro. Mixing of P1 and P2 formed the P1-P2 heterodimer, as demonstrated by gel mobility shift and chemical crosslinking. This heterodimer, but neither P1 or P2 alone, tightly bound to P0 and formed a pentameric complex, presumably as P0(P1-P2)2, assumed from its molecular weight derived from sedimentation analysis. Complex formation strongly stimulated binding of P0 to the GTPase-associated RNA domain. The protein complex and eL12 (E.coli L11-type), which cross-bound to the E.coli equivalent RNA domain, were tested for their function by replacing with the E.coli counterparts L10.L7/L12 complex and L11 on the rRNA domain within the 50S subunits. Both P1 and P2, together with P0 and eL12, were required to activate ribosomes in polyphenylalanine synthesis dependent on eucaryotic elongation factors as well as eEF-2-dependent GTPase activity. The results suggest that formation of the P1-P2 heterodimer is required for subsequent formation of the P0(P1-P2)2 complex and its functional rRNA binding in silkworm ribosomes.
Shimizu
Tomomi
T
Institute of High Polymer Research and Department of Applied Biological Science, Faculty of Textile Science and Technology, Shinshu University, Ueda 386-8567, Japan.
Nakagaki
Masao
M
Nishi
Yoshinori
Y
Kobayashi
Yuji
Y
Hachimori
Akira
A
Uchiumi
Toshio
T
D000595:Amino Acid Sequence; D000818:Animals; D012831:Bombyx; D019281:Dimerization; D020558:GTP Phosphohydrolases / Q000737:chemistry; D019476:Insect Proteins / Q000378:metabolism*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012339:RNA, Ribosomal, 28S / Q000737:chemistry* / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism; D012269:Ribosomal Proteins / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D019476:Insect Proteins; D046911:Macromolecular Substances; D010750:Phosphoproteins; D012339:RNA, Ribosomal, 28S; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins; C031937:phosphoprotein P2, ribosomal; C035690:ribosomal phosphoprotein P1; C054675:ribosomal protein P0; D020558:GTP Phosphohydrolases