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Title: Designing seryl‐ tRNA synthetase for improved serylation of selenocysteine tRNA s

Abstract

Selenocysteine (Sec) lacks a cognate aminoacyl‐ tRNA synthetase. Instead, seryl‐ tRNA synthetase (Ser RS ) produces Ser‐ tRNA S ec , which is subsequently converted by selenocysteine synthase to Sec‐ tRNA S ec . Escherichia coli Ser RS serylates tRNA S ec poorly; this may hinder efficient production of designer selenoproteins in vivo . Guided by structural modelling and selection for chloramphenicol acetyltransferase activity, we evolved three Ser RS variants capable of improved Ser‐ tRNA S ec synthesis. They display 10‐, 8‐, and 4‐fold increased k cat / K M values compared to wild‐type Ser RS using synthetic tRNA S ec species as substrates. The enzyme variants also facilitate in vivo read‐through of a UAG codon in the position of the critical serine 146 of chloramphenicol acetyltransferase. These results indicate that the naturally evolved Ser RS is capable of further evolution for increased recognition of a specific tRNA isoacceptor.

Authors:
 [1]; ORCiD logo [1]; ORCiD logo [1]; ORCiD logo [2]
  1. Department of Molecular Biophysics &, Biochemistry Yale University New Haven CT USA
  2. Department of Molecular Biophysics &, Biochemistry Yale University New Haven CT USA, Department of Chemistry Yale University New Haven CT USA
Publication Date:
Sponsoring Org.:
USDOE
OSTI Identifier:
1478680
Resource Type:
Publisher's Accepted Manuscript
Journal Name:
FEBS Letters
Additional Journal Information:
Journal Name: FEBS Letters Journal Volume: 592 Journal Issue: 22; Journal ID: ISSN 0014-5793
Publisher:
Wiley Blackwell (John Wiley & Sons)
Country of Publication:
Netherlands
Language:
English

Citation Formats

Fu, Xian, Crnković, Ana, Sevostyanova, Anastasia, and Söll, Dieter. Designing seryl‐ tRNA synthetase for improved serylation of selenocysteine tRNA s. Netherlands: N. p., 2018. Web. doi:10.1002/1873-3468.13271.
Fu, Xian, Crnković, Ana, Sevostyanova, Anastasia, & Söll, Dieter. Designing seryl‐ tRNA synthetase for improved serylation of selenocysteine tRNA s. Netherlands. https://doi.org/10.1002/1873-3468.13271
Fu, Xian, Crnković, Ana, Sevostyanova, Anastasia, and Söll, Dieter. Wed . "Designing seryl‐ tRNA synthetase for improved serylation of selenocysteine tRNA s". Netherlands. https://doi.org/10.1002/1873-3468.13271.
@article{osti_1478680,
title = {Designing seryl‐ tRNA synthetase for improved serylation of selenocysteine tRNA s},
author = {Fu, Xian and Crnković, Ana and Sevostyanova, Anastasia and Söll, Dieter},
abstractNote = {Selenocysteine (Sec) lacks a cognate aminoacyl‐ tRNA synthetase. Instead, seryl‐ tRNA synthetase (Ser RS ) produces Ser‐ tRNA S ec , which is subsequently converted by selenocysteine synthase to Sec‐ tRNA S ec . Escherichia coli Ser RS serylates tRNA S ec poorly; this may hinder efficient production of designer selenoproteins in vivo . Guided by structural modelling and selection for chloramphenicol acetyltransferase activity, we evolved three Ser RS variants capable of improved Ser‐ tRNA S ec synthesis. They display 10‐, 8‐, and 4‐fold increased k cat / K M values compared to wild‐type Ser RS using synthetic tRNA S ec species as substrates. The enzyme variants also facilitate in vivo read‐through of a UAG codon in the position of the critical serine 146 of chloramphenicol acetyltransferase. These results indicate that the naturally evolved Ser RS is capable of further evolution for increased recognition of a specific tRNA isoacceptor.},
doi = {10.1002/1873-3468.13271},
journal = {FEBS Letters},
number = 22,
volume = 592,
place = {Netherlands},
year = {2018},
month = {10}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
https://doi.org/10.1002/1873-3468.13271

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Cited by: 5 works
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