Crystal structure of the flavin-dependent thymidylate synthase Thy1 from Thermus thermophilus with an extra C-terminal domain

The thymidylate synthases ThyA and Thy1 are enzymes that catalyse the formation of thymidine monophosphate from 2′-deoxyuridine monophosphate. Thy1 (or ThyX) requires flavin for catalytic reactions, while ThyA does not. In the present study, the crystal structure of the flavin-dependent thymidylate synthase Thy1 from Thermus thermophilus HB8 (TtThy1, TTHA1096) was determined in complex with FAD and phosphate at 2.5 Å resolution. TtThy1 is a tetrameric molecule like other Thy1 proteins, to which four FAD molecules are bound. In the crystal of TtThy1, two phosphate ions were bound to each dUMP-binding site. The characteristic feature of TtThy1 is the existence of an extra C-terminal domain (CTD) consisting of three α-helices and a β-strand. The function of the CTD is unknown and database analysis showed that this CTD is only shared by part of the Deinococcus–Thermus phylum.

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Crystal Structure and Nucleotide Binding of the Thermus thermophilus RNA Helicase Hera N-terminal Domain

Journal of Molecular Biology ◽

10.1016/j.jmb.2006.06.065 ◽

2006 ◽

Vol 361 (4) ◽

pp. 731-743 ◽

Cited By ~ 26

Author(s):

Markus G. Rudolph ◽

Ramona Heissmann ◽

Julia G. Wittmann ◽

Dagmar Klostermeier

Keyword(s):

Crystal Structure ◽

Thermus Thermophilus ◽

Nucleotide Binding ◽

Rna Helicase ◽

Terminal Domain

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Crystal structure studies of NADP+ dependent isocitrate dehydrogenase from Thermus thermophilus exhibiting a novel terminal domain

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2014.04.164 ◽

2014 ◽

Vol 449 (1) ◽

pp. 107-113 ◽

Cited By ~ 3

Author(s):

S.M. Kumar ◽

K.J. Pampa ◽

M. Manjula ◽

M.M.M. Abdoh ◽

Naoki Kunishima ◽

...

Keyword(s):

Crystal Structure ◽

Isocitrate Dehydrogenase ◽

Thermus Thermophilus ◽

Terminal Domain

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Mechanistic and structural basis for inhibition of thymidylate synthase ThyX

Open Biology ◽

10.1098/rsob.120120 ◽

2012 ◽

Vol 2 (10) ◽

pp. 120120 ◽

Cited By ~ 22

Author(s):

Tamara Basta ◽

Yap Boum ◽

Julien Briffotaux ◽

Hubert F. Becker ◽

Isabelle Lamarre-Jouenne ◽

...

Keyword(s):

Crystal Structure ◽

Thymidylate Synthase ◽

Active Site ◽

De Novo ◽

Tight Binding ◽

Binding Pocket ◽

Structural Basis ◽

Microbial Genomes ◽

Thymidylate Synthases ◽

Thymidylate Synthesis

Nature has established two mechanistically and structurally unrelated families of thymidylate synthases that produce de novo thymidylate or dTMP, an essential DNA precursor. Representatives of the alternative flavin-dependent thymidylate synthase family, ThyX, are found in a large number of microbial genomes, but are absent in humans. We have exploited the nucleotide binding pocket of ThyX proteins to identify non-substrate-based tight-binding ThyX inhibitors that inhibited growth of genetically modified Escherichia coli cells dependent on thyX in a manner mimicking a genetic knockout of thymidylate synthase. We also solved the crystal structure of a viral ThyX bound to 2-hydroxy-3-(4-methoxybenzyl)-1,4-naphthoquinone at a resolution of 2.6 Å. This inhibitor was found to bind within the conserved active site of the tetrameric ThyX enzyme, at the interface of two monomers, partially overlapping with the dUMP binding pocket. Our studies provide new chemical tools for investigating the ThyX reaction mechanism and establish a novel mechanistic and structural basis for inhibition of thymidylate synthesis. As essential ThyX proteins are found e.g. in Mycobacterium tuberculosis and Helicobacter pylori , our studies have also potential to pave the way towards the development of new anti-microbial compounds.

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