Xanthine Derivatives Reveal an Allosteric Binding Site in Methylenetetrahydrofolate Dehydrogenase 2 (MTHFD2).

Lung-Chun Lee,Hsin-Huei Chang,Chih-Hsiang Huang,Su-Ying Wu,Tsu Hsu,Ching-Cheng Hsueh,Fang-Chun Kung,Weir-Torn Jiaang,Yi-Hui Peng,Ching-Chuan Kuo,Cheng-Tai Lu

doi:10.1021/acs.jmedchem.1c00663

Abstract

Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) plays an important role in one-carbon metabolism. The MTHFD2 gene is upregulated in various cancers but very low or undetectable in normal proliferating cells, and therefore a potential target for cancer treatment. In this study, we present the structure of MTHFD2 in complex with xanthine derivative 15, which allosterically binds to MTHFD2 and coexists with the substrate analogue. A kinetic study demonstrated the uncompetitive inhibition of MTHFD2 by 15. Allosteric inhibitors often provide good selectivity and, indeed, xanthine derivatives are highly selective for MTHFD2. Moreover, several conformational changes were observed upon the binding of 15, which impeded the binding of the cofactor and phosphate to MTHFD2. To the best of our knowledge, this is the first study to identify allosteric inhibitors targeting the MTHFD family and our results would provide insights on the inhibition mechanism of MTHFD proteins and the development of novel inhibitors.

Highlights

Cellular metabolism is substantially altered during tumorigenesis and malignant progression
Depletion of Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) impairs aggressive phenotypes and causes cell death in multiple cancers, including breast cancer, colorectal cancer, central nervous system tumors, lung cancer, ovarian cancer, renal cancer, melanoma, and leukemia.[6−8] These results suggested that targeting MTHFD2 is a promising strategy for cancer therapy.[9,10]
The mechanistic basis for the allosteric inhibition of MTHFD2 by xanthine derivative 15 has been elucidated based on the inhibitor-bound MTHFD2 structure, which was solved at a resolution of 2.13 Å

Summary

■ INTRODUCTION

Cellular metabolism is substantially altered during tumorigenesis and malignant progression. Arg[142] could no longer form the hydrogen bond with 3 and the flexible side chain of Arg[142] shifted away (Figure 4A) All of these structural differences as compared with MTHFD2/3/21 and MTHFD2/ 15/21 explain the decreased inhibitory activity of 3. Arg[201], located in the βe-αE loop, plays an important role in phosphate binding and contributes to the activity of human MTHFD2 as the mutation of Arg[201] to lysine resulted in the total loss of enzymatic activity.[14] In the presence of NAD+, Arg[201] serves as the interaction center to form hydrogen bonds with the phosphate ion, the adenosine ribose ring of NAD+, and coordinates with Asp216′, His219′, and Asp225′ in monomer B through the salt bridge and hydrogen-bond network (Figure 6B,C). In addition to contributing to the dimer interactions, these residues play roles in maintaining the activity of MTHFD2

■ DISCUSSION AND CONCLUSIONS

■ ACKNOWLEDGMENTS

■ REFERENCES