Abstract
Human thymidylate synthase (hTS) is a 72 kDa homodimeric enzyme responsible for the conversion of deoxyuridine monophosphate (dUMP) to deoxythymidine monophosphate (dTMP), making it the sole source of de novo dTMP in human cells. As a result, hTS is an attractive anti-cancer therapeutic target. Additionally, hTS is known to possess a number of interesting biophysical features, including adoption of active and inactive conformations, positively cooperative substrate binding, half-the-sites activity, and interacting with its own mRNA. The physical mechanisms underlying these properties, and how they may be leveraged to guide therapeutic development, are yet to be fully explored. Here, as a preface to detailed NMR characterization, we present backbone amide and ILVM methyl resonance assignments for hTS in apo and dUMP bound forms. In addition, we present backbone amide resonance assignments for hTS bound to a substrate analog and the native cofactor.
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Availability of data and material
The chemical shift assignments are deposited in the Biological Magnetic Resonance Data Bank under accession numbers 50520, 50521, and 50519 for apo, dUMP, and diligand bound states. The NMRViewJ software is available at https://nmrfx.org.
Code availability
Files needed for MAGIC analysis of NOESY data are available at https://github.com/NMRsoftware/MAGIC.
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Acknowledgements
We thank Dr. Amnon Kohen for providing a vector containing the hTS coding sequence. We also thank Dr. Gregory Young and Dr. Stewart Parnham of the UNC Biomolecular NMR Facility for assistance in NMR data collection.
Funding
This work was funded by National Institutes of Health (NIH) grant GM083059 to A.L.L and NIH Grant 5 T32 GM 8570-22 to J.P.B. This work was supported in part by the National Cancer Institute of the NIH under Award Number P30CA016086 Cancer Center Support Grant. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.
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ALL and JPB: Research was designed; JPB performed and analyzed NMR experiments. JPB. and ALL wrote the manuscript.
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Bonin, J.P., Lee, A.L. Backbone and ILVM methyl resonance assignments of human thymidylate synthase in apo and substrate bound forms. Biomol NMR Assign 15, 197–202 (2021). https://doi.org/10.1007/s12104-021-10006-x
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DOI: https://doi.org/10.1007/s12104-021-10006-x