Discovery of natural CdnP inhibitors through structure-based virtual screening and molecular dynamics simulations
Xiaoxia Gu1,2, Chaohu Xiong2, Xinyu Wang2
1Department of Pharmacy, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
Abstract:
Tuberculosis, caused by Mycobacterium tuberculosis, remains one of the most lethal infectious diseases both historically and in the post-coronavirus disease 2019 era. CdnP (Rv2837c) functions as a bifunctional oligoribonuclease and 3'(2')-phosphoadenosine 5'-phosphate-phosphatase that impedes host immune responses by recognizing bacterial cyclic dinucleotides such as c-di-AMP, which serve as pathogen-associated molecular patterns. Despite its significance, strategies targeting CdnP remain limited. Through high-throughput virtual screening and enzymatic assays, we identified four natural product inhibitors: one coumarin derivative (macrosporusone A) and three flavonoid glucosides (ligustroflavone, rhoifolin, and neodiosmin). Surface plasmon resonance measurements confirmed direct binding of these compounds to CdnP with nanomolar to micromolar affinities. Molecular dynamics simulations elucidated a dual inhibitory mechanism wherein these compounds competitively occupy the product (AMP)-binding site while simultaneously constraining conformational plasticity of the substrate-binding domain. Evolutionary analysis demonstrated that these inhibitors exhibit broad-spectrum activity against bacterial CdnP orthologs while showing minimal inhibition of host-derived 2',3'-cGAMP-specific phosphodiesterases, suggesting favorable selectivity. Notably, ligustroflavone exhibited superior inhibitory potency. In contrast, FDA-approved phosphodiesterase inhibitors showed poor activity against bacterial orthologs. These findings provide a foundation for developing novel host-directed therapeutics against tuberculosis that could potentially enhance stimulator of interferon genes (STING)-mediated immune responses without exerting selective pressure for antimicrobial resistance.
Importance:
Tuberculosis (TB) remains a leading cause of mortality worldwide, with drug resistance posing a significant challenge to global control efforts. This study represents a major contribution to the field by identifying novel natural product inhibitors targeting CdnP (Rv2837c), a c-di-AMP-specific phosphodiesterase critical for Mycobacterium tuberculosis pathogenesis. The significance of this work lies in its innovative approach to TB therapy by perturbing bacterial nucleotide signaling pathways rather than directly inhibiting bacterial growth. By selectively targeting bacterial CdnP while avoiding host phosphodiesterases, these compounds-particularly ligustroflavone and other flavonoid glucosides-offer a promising foundation for developing host-directed therapeutics with potentially reduced selective pressure for antimicrobial resistance. Furthermore, the detailed structural insights and inhibitory mechanisms elucidated through molecular dynamics simulations provide valuable knowledge for rational drug design. This research bridges natural product discovery with computational biology to address the urgent need for novel TB treatments, especially against drug-resistant strains, presenting a significant advancement toward more effective therapeutic interventions for this persistent global health threat.
Insights
Researchers discovered natural compounds that inhibit CdnP, an enzyme crucial for tuberculosis bacteria. These inhibitors offer a new strategy for tuberculosis treatment, potentially overcoming drug resistance and enhancing immune responses.
Area of Science:
- Biochemistry and Molecular Biology
- Natural Product Drug Discovery
- Computational Biology
Background:
- Tuberculosis (TB), caused by Mycobacterium tuberculosis, remains a major global health threat, exacerbated by increasing drug resistance.
- The bacterial enzyme CdnP (Rv2837c) is essential for TB pathogenesis by modulating host immune responses via cyclic dinucleotides.
- Targeting CdnP presents a novel therapeutic strategy distinct from traditional growth inhibition, potentially circumventing existing resistance mechanisms.
Purpose of the Study:
- To identify novel natural product inhibitors of Mycobacterium tuberculosis CdnP.
- To elucidate the inhibitory mechanism and assess the selectivity of identified compounds.
- To provide a foundation for developing new host-directed therapeutics against TB.
Main Methods:
- High-throughput virtual screening and enzymatic assays to identify potential inhibitors.
- Surface plasmon resonance (SPR) for direct binding affinity measurements.
- Molecular dynamics (MD) simulations to understand inhibitory mechanisms and evolutionary analysis for ortholog selectivity.
Main Results:
- Four natural products, including macrosporusone A (coumarin) and three flavonoid glucosides (ligustroflavone, rhoifolin, neodiosmin), were identified as CdnP inhibitors.
- These compounds bind directly to CdnP with nanomolar to micromolar affinities, exhibiting a dual inhibitory mechanism at the AMP-binding site and substrate-binding domain.
- Inhibitors showed broad-spectrum activity against bacterial CdnP orthologs with high selectivity over human phosphodiesterases; ligustroflavone demonstrated superior potency.
Conclusions:
- Novel natural product inhibitors targeting bacterial CdnP have been discovered, offering a promising avenue for TB drug development.
- The identified compounds, particularly flavonoid glucosides, provide a basis for rational drug design of host-directed TB therapeutics.
- This approach may enhance STING-mediated immunity and reduce selective pressure for antimicrobial resistance, addressing critical needs in TB control.
