Functional involvement of a conserved motif in the middle region of the human ribosomal protein eL42 in translation

Ribosomal protein eL42 (formerly known as L36A), a small protein of the large (60S) subunit of the eukaryotic ribosome, is a component of its exit (E) site. The residue K53 of this protein resides within the motif QSGYGGQTK mainly conserved in eukaryotes, and it is located in the immediate vicinity...

Full description

Saved in:
Bibliographic Details
Published in:Biochimie 2024-03, Vol.218, p.96-104
Main Authors: Bulygin, Konstantin N., Malygin, Alexey A., Graifer, Dmitri M.
Format: Article
Language:English
Subjects:
Amino Acids - metabolism
Elongation phase of protein synthesis
HEK293 Cells
HEK293T cells
Humans
Polysome profiling
Protein Biosynthesis
Ribosomal protein eL42
Ribosomal Proteins - metabolism
Ribosomes - metabolism
RNA, Transfer - metabolism
Transient transfection
Citations: Items that this one cites
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Ribosomal protein eL42 (formerly known as L36A), a small protein of the large (60S) subunit of the eukaryotic ribosome, is a component of its exit (E) site. The residue K53 of this protein resides within the motif QSGYGGQTK mainly conserved in eukaryotes, and it is located in the immediate vicinity of the CCA-terminus of the ribosome-bound tRNA in the hybrid P/E state. To examine the role of this eL42 motif in translation, we obtained HEK293T cells producing the wild-type FLAG-tagged protein or its mutant forms with either single substitutions of conserved amino acid residues in the above motif, or simultaneous replacements in positions 45 and 51 or 45 and 53. Examination of the level of exogenous eL42 in fractions of polysome profiles from the target protein-producing cells by the Western blotting revealed that neither single substitution affects the assembly of 60S ribosomal subunits and 80S ribosomes or critically decreases the level of polysomes, but the latter was observed with the double replacements. Analysis of tRNAs bound to 80S ribosomes containing eL42 with double substitutions and examination their peptidyl transferase activity enabled estimation the stage of the elongation cycle, in which amino acid residues of the conserved eL42 motif are involved. We clearly show that cooperative interactions implicating the eL42 residues Q45, Q51, and K53 play a critical role in the ability of the human ribosome to perform properly elongation cycle at the step of deacylated tRNA dissociation from the E site in the human cell. •Conserved region 45–53 of RP eL42 is crucial for translation elongation in human cells.•Double replacements of amino acid residues in this eL42 region blocks elongation.•The role of the 45–53 region is providing proper tRNA dissociation from the E site.
ISSN:0300-9084
1638-6183
DOI:10.1016/j.biochi.2023.09.010