bi9b00528_si_001.pdf (30.55 MB)
Structural and Mutagenesis Studies Evince the Role of the Extended Protuberant Domain of Ribosomal Protein uL10 in Protein Translation
journal contribution
posted on 2019-08-22, 19:04 authored by Kwok-Ho
Andrew Choi, Lei Yang, Ka-Ming Lee, Conny Wing-Heng Yu, David K. Banfield, Kosuke Ito, Toshio Uchiumi, Kam-Bo WongThe
lateral stalk of ribosomes constitutes the GTPase-associated
center and is responsible for recruiting translation factors to the
ribosomes. The eukaryotic stalk contains a P-complex, in which one
molecule of uL10 (formerly known as P0) protein binds two copies of
P1/P2 heterodimers. Unlike bacterial uL10, eukaryotic uL10 has an
extended protuberant (uL10ext) domain inserted into the N-terminal
RNA-binding domain. Here, we determined the solution structure of
the extended protuberant domain of Bombyx mori uL10
by nuclear magnetic resonance spectroscopy. Comparison of the structures
of the B. mori uL10ext domain with eRF1-bound and
eEF2-bound ribosomes revealed significant structural rearrangement
in a “hinge” region surrounding Phe183, a residue conserved
in eukaryotic but not in archaeal uL10. 15N relaxation
analyses showed that residues in the hinge region have significantly
large values of transverse relaxation rates. To test the role of the
conserved phenylalanine residue, we created a yeast mutant strain
expressing an F181A variant of uL10. An in vitro translation
assay showed that the alanine substitution increased the level of
polyphenylalanine synthesis by ∼33%. Taken together, our results
suggest that the hinge motion of the uL10ext domain facilitates the
binding of different translation factors to the GTPase-associated
center during protein synthesis.