Mechanistic dissection of premature translation termination induced by acidic residues-enriched nascent peptide

Chadani, Yuhei; Kanamori, Takashi; Niwa, Tatsuya; Ichihara, Kazuya; Nakayama, Keiichi I.; Matsumoto, Akinobu; Taguchi, Hideki

doi:10.1016/j.celrep.2023.113569

Cited by 2 publications

(1 citation statement)

References 62 publications

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“…The major advantage of the system is that it consists only of purified factors that are required for translation, and it is devoid of nucleases, proteases, and other unrelated factors, such as metabolic enzymes [1]. The PURE system is currently used not only for studying the mechanism of translation and protein folding [2][3][4][5][6][7][8], but also in synthetic biology and protein engineering applications [9][10][11][12][13][14][15][16]. The yield of the proteins synthesized using the PURE system was initially lower than that of the E. coli S30 extract system, but it has now increased to 1 mg/mL following optimization of the reaction composition [17].…”

Section: Introductionmentioning

confidence: 99%

N-Terminal Amino Acid Affects the Translation Efficiency at Lower Temperatures in a Reconstituted Protein Synthesis System

Fuse-Murakami,

Matsumoto,

Kanamori

2024

IJMS

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The Escherichia coli (E. coli)-based protein synthesis using recombinant elements (PURE) system is a cell-free protein synthesis system reconstituted from purified factors essential for E. coli translation. The PURE system is widely used for basic and synthetic biology applications. One of the major challenges associated with the PURE system is that the protein yield of the system varies depending on the protein. Studies have reported that the efficiency of translation is significantly affected by nucleotide and amino acid sequences, especially in the N-terminal region. Here, we investigated the inherent effect of various N-terminal sequences on protein synthesis using the PURE system. We found that a single amino acid substitution in the N-terminal region significantly altered translation efficiency in the PURE system, especially at low temperatures. This result gives us useful suggestions for the expression of the protein of interest in vitro and in vivo.

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Section: Introductionmentioning

confidence: 99%

N-Terminal Amino Acid Affects the Translation Efficiency at Lower Temperatures in a Reconstituted Protein Synthesis System

Fuse-Murakami,

Matsumoto,

Kanamori

2024

IJMS

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The ABCF proteins in Escherichia coli individually cope with ‘hard-to-translate’ nascent peptide sequences

Chadani,

Yamanouchi,

Uemura

et al. 2024

Nucleic Acids Research

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Organisms possess a wide variety of proteins with diverse amino acid sequences, and their synthesis relies on the ribosome. Empirical observations have led to the misconception that ribosomes are robust protein factories, but in reality, they have several weaknesses. For instance, ribosomes stall during the translation of the proline-rich sequences, but the elongation factor EF-P assists in synthesizing proteins containing the poly-proline sequences. Thus, living organisms have evolved to expand the translation capability of ribosomes through the acquisition of translation elongation factors. In this study, we have revealed that Escherichia coli ATP-Binding Cassette family-F (ABCF) proteins, YheS, YbiT, EttA and Uup, individually cope with various problematic nascent peptide sequences within the exit tunnel. The correspondence between noncanonical translations and ABCFs was YheS for the translational arrest by nascent SecM, YbiT for poly-basic sequence-dependent stalling and poly-acidic sequence-dependent intrinsic ribosome destabilization (IRD), EttA for IRD at the early stage of elongation, and Uup for poly-proline-dependent stalling. Our results suggest that ATP hydrolysis-coupled structural rearrangement and the interdomain linker sequence are pivotal for handling ‘hard-to-translate’ nascent peptides. Our study highlights a new aspect of ABCF proteins to reduce the potential risks that are encoded within the nascent peptide sequences.

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Mechanistic dissection of premature translation termination induced by acidic residues-enriched nascent peptide

Cited by 2 publications

References 62 publications

N-Terminal Amino Acid Affects the Translation Efficiency at Lower Temperatures in a Reconstituted Protein Synthesis System

N-Terminal Amino Acid Affects the Translation Efficiency at Lower Temperatures in a Reconstituted Protein Synthesis System

The ABCF proteins in Escherichia coli individually cope with ‘hard-to-translate’ nascent peptide sequences

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