Crystal structure and enzymatic characterization of thymidylate synthase X from Helicobacter pylori strain SS1 |
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Authors: | Wang Kuifeng Wang Qi Chen Jing Chen Lili Jiang Hualiang Shen Xu |
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Affiliation: | State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China. |
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Abstract: | Thymidylate synthase X (ThyX) catalyzes the methylation of dUMP to form dTMP in bacterial life cycle and is regarded as a promising target for antibiotics discovery. Helicobacter pylori is a human pathogen associated with a number of human diseases. Here, we cloned and purified the ThyX enzyme from H. pylori SS1 strain (HpThyX). The recombinant HpThyX was discovered to exhibit the maximum activity at pH 8.5, and Km values of the two substrates dUMP and CH2H4folate were determined to be 15.3 ± 1.25 μM and 0.35 ± 0.18 mM, respectively. The analyzed crystal structure of HpThyX with the cofactor FAD and the substrate dUMP (at 2.31 Å) revealed that the enzyme was a tetramer bound to four dUMP and four FAD molecules. Different from the catalytic feature of the classical thymidylate synthase (ThyA), N5 atom of the FAD functioned as a nucleophile in the catalytic reaction instead of Ser84 and Ser85 residues. Our current work is expected to help better understand the structural and enzymatic features of HpThyX thus further providing valuable information for anti‐H. pylori inhibitor discovery. |
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Keywords: | thymidylate synthase Helicobacter pylori thymidylate synthase ThyA |
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