Research output: Contribution to journal › Article › peer-review
The functional role of the C-terminal tail of the human ribosomal protein uS19. / Bulygin, Konstantin; Malygin, Alexey; Gopanenko, Alexander et al.
In: Biochimica et Biophysica Acta - Gene Regulatory Mechanisms, Vol. 1863, No. 3, 194490, 03.2020.Research output: Contribution to journal › Article › peer-review
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TY - JOUR
T1 - The functional role of the C-terminal tail of the human ribosomal protein uS19
AU - Bulygin, Konstantin
AU - Malygin, Alexey
AU - Gopanenko, Alexander
AU - Graifer, Dmitri
AU - Karpova, Galina
N1 - Publisher Copyright: © 2020 Elsevier B.V.
PY - 2020/3
Y1 - 2020/3
N2 - The eukaryotic ribosomal protein uS19 has a C-terminal tail that is absent in its bacterial homologue. This tail has been shown to be involved in the formation of the decoding site of human ribosomes. We studied here the previously unexplored functional significance of the 15 C-terminal amino acid residues of human uS19 for the assembly of ribosomes and translation using HEK293-based cell cultures capable of producing FLAG-labeled uS19 (uS19FLAG) or its mutant form deprived of the mentioned amino acid ones. The examination of polysome profiles of cytoplasmic extracts from the respective cells revealed that the deletion of the above uS19 amino acid residues barely affected the assembly and maturation of 40S subunits and the initiation of translation, but completely prevented the formation of polysomes. This implied the crucial importance of the uS19 tail in the elongation process. Analysis of tRNAs associated with 40S subunits and 80S ribosomes containing wild type uS19FLAG or its truncated form showed that the deletion of the C-terminal pentadecapeptide fragment of uS19 did not interfere with the binding of aminoacyl-tRNA (aa-tRNA) at the ribosomal A site. The results led to the conclusion that the transpeptidation, which occurs on the large ribosomal subunit after decoding the A site codon by the incoming aa-tRNA, is the most likely elongation stage, where this uS19 fragment can play a critical role. Our findings suggest that the uS19 tail is a keystone player in the accommodation of aa-tRNA at the A site, which is a pre-requisite for the peptide transfer.
AB - The eukaryotic ribosomal protein uS19 has a C-terminal tail that is absent in its bacterial homologue. This tail has been shown to be involved in the formation of the decoding site of human ribosomes. We studied here the previously unexplored functional significance of the 15 C-terminal amino acid residues of human uS19 for the assembly of ribosomes and translation using HEK293-based cell cultures capable of producing FLAG-labeled uS19 (uS19FLAG) or its mutant form deprived of the mentioned amino acid ones. The examination of polysome profiles of cytoplasmic extracts from the respective cells revealed that the deletion of the above uS19 amino acid residues barely affected the assembly and maturation of 40S subunits and the initiation of translation, but completely prevented the formation of polysomes. This implied the crucial importance of the uS19 tail in the elongation process. Analysis of tRNAs associated with 40S subunits and 80S ribosomes containing wild type uS19FLAG or its truncated form showed that the deletion of the C-terminal pentadecapeptide fragment of uS19 did not interfere with the binding of aminoacyl-tRNA (aa-tRNA) at the ribosomal A site. The results led to the conclusion that the transpeptidation, which occurs on the large ribosomal subunit after decoding the A site codon by the incoming aa-tRNA, is the most likely elongation stage, where this uS19 fragment can play a critical role. Our findings suggest that the uS19 tail is a keystone player in the accommodation of aa-tRNA at the A site, which is a pre-requisite for the peptide transfer.
KW - Aminoacyl-tRNA accommodation
KW - Decoding site
KW - Deletion of uS19 C-terminal amino acid residues
KW - Human ribosomal protein uS19
KW - Small ribosomal subunit
KW - Translation elongation
KW - CONTACTS
KW - SITE
KW - STATES
KW - CRYSTAL-STRUCTURE
KW - STOP CODONS
KW - RELEASE FACTOR ERF1
KW - ELEMENTS
KW - MESSENGER-RNA
KW - ELONGATION
KW - SUBUNIT
UR - http://www.scopus.com/inward/record.url?scp=85078722479&partnerID=8YFLogxK
U2 - 10.1016/j.bbagrm.2020.194490
DO - 10.1016/j.bbagrm.2020.194490
M3 - Article
C2 - 31991215
AN - SCOPUS:85078722479
VL - 1863
JO - Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
JF - Biochimica et Biophysica Acta - Gene Regulatory Mechanisms
SN - 1874-9399
IS - 3
M1 - 194490
ER -
ID: 23329115