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Crystallographic study of the 2‐thioribothymidine‐synthetic complex TtuA–TtuB from Thermus thermophilus
Author(s) -
Chen Minghao,
Narai Shun,
Omura Naoki,
Shigi Naoki,
Chimnaronk Sarin,
Tanaka Yoshikazu,
Yao Min
Publication year - 2016
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x16014242
Subject(s) - thermus thermophilus , thermophile , chemistry , covalent bond , crystallography , residue (chemistry) , stereochemistry , crystal structure , transferase , biochemistry , escherichia coli , enzyme , organic chemistry , gene
The ubiquitin‐like protein TtuB is a sulfur carrier for the biosynthesis of 2‐thioribothymidine (s 2 T) at position 54 in some thermophilic bacterial tRNAs. TtuB captures a S atom at its C‐terminus as a thiocarboxylate and transfers it to tRNA by the transferase activity of TtuA. TtuB also functions to suppress s 2 T formation by forming a covalent bond with TtuA. To explore how TtuB interacts with TtuA and switches between these two different functions, high‐resolution structure analysis of the TtuA–TtuB complex is required. In this study, the TtuA–TtuB complex from Thermus thermophilus was expressed, purified and crystallized. To mimic the thiocarboxylated TtuB, the C‐terminal Gly residue was replaced with Cys (G65C) to obtain crystals of the TtuA–TtuB complex. A Zn‐MAD data set was collected to a resolution of 2.5 Å. MAD analysis successfully determined eight Zn sites, and a partial structure model composed of four TtuA–TtuB complexes in the asymmetric unit was constructed.