Filamentation and inhibition of prokaryotic CTP synthase with ligands

  • Chenjun Guo 1 ,
  • Zixuan Wang 1 ,
  • Ji-Long Liu , 1,2,3
Expand
  • 1. School of Life Science and Technology, ShanghaiTech University, Shanghai, China.
  • 2. Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK.
  • 3. Shanghai Clinical Research and Trial Center, Shanghai, China.
liujl3@shanghaitech.edu.cn; jilong.liu@dpag.ox.ac.uk

Received date: 09 Nov 2023

Accepted date: 06 Feb 2024

Published date: 20 Feb 2024

Copyright

2024 2024 The Authors. mLife published by John Wiley & Sons Australia, Ltd on behalf of Institute of Microbiology, Chinese Academy of Sciences.

Abstract

Cytidine triphosphate synthase (CTPS) plays a pivotal role in the de novo synthesis of cytidine triphosphate (CTP), a fundamental building block for RNA and DNA that is essential for life. CTPS is capable of directly binding to all four nucleotide triphosphates: adenine triphosphate, uridine triphosphate, CTP, and guanidine triphosphate. Furthermore, CTPS can form cytoophidia in vivo and metabolic filaments in vitro, undergoing regulation at multiple levels. CTPS is considered a potential therapeutic target for combating invasions or infections by viral or prokaryotic pathogens. Utilizing cryo-electron microscopy, we determined the structure of Escherichia coli CTPS (ecCTPS) filament in complex with CTP, nicotinamide adenine dinucleotide (NADH), and the covalent inhibitor 6-diazo-5-oxo-L-norleucine (DON), achieving a resolution of 2.9 Å. We constructed a phylogenetic tree based on differences in filament-forming interfaces and designed a variant to validate our hypothesis, providing an evolutionary perspective on CTPS filament formation. Our computational analysis revealed a solvent-accessible ammonia tunnel upon DON binding. Through comparative structural analysis, we discern a distinct mode of CTP binding of ecCTPS that differs from eukaryotic counterparts. Combining biochemical assays and structural analysis, we determined and validated the synergistic inhibitory effects of CTP with NADH or adenine on CTPS. Our results expand our comprehension of the diverse regulatory aspects of CTPS and lay a foundation for the design of specific inhibitors targeting prokaryotic CTPS.

Cite this article

Chenjun Guo , Zixuan Wang , Ji-Long Liu . Filamentation and inhibition of prokaryotic CTP synthase with ligands[J]. mLife, 2024 , 3(2) : 240 -250 . DOI: 10.1002/mlf2.12119

Options
Outlines

/