Tipiracil binds to uridine site and inhibits Nsp15 endoribonuclease NendoU from SARS-CoV-2

• Published in: Communications biology 2021-02, Vol.4 (1), p.193-11, Article 193
• Main Authors: Kim, Youngchang, Wower, Jacek, Maltseva, Natalia, Chang, Changsoo, Jedrzejczak, Robert, Wilamowski, Mateusz, Kang, Soowon, Nicolaescu, Vlad, Randall, Glenn, Michalska, Karolina, Joachimiak, Andrzej
• Format: Article
• Language: English
• Subjects: 45/29; 631/337; 631/535; 82/16; 82/80; 82/83; A549 Cells; Antiviral Agents - chemistry; Antiviral Agents - pharmacokinetics; Antiviral Agents - pharmacology; BASIC BIOLOGICAL SCIENCES; Binding sites; Biology; Biomedical and Life Sciences; Catalytic Domain; Colorectal carcinoma; Coronaviruses; COVID-19; COVID-19 - drug therapy; COVID-19 - virology; Crystallography; Crystallography, X-Ray; Drug development; Endoribonucleases - antagonists & inhibitors; Endoribonucleases - chemistry; Endoribonucleases - metabolism; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - pharmacokinetics; Enzyme Inhibitors - pharmacology; Enzymes; Humans; Immune response; Immunosuppressive agents; Innate immunity; Life Sciences; Ligands; Manganese; Metal ions; Models, Molecular; Molecular biology; Nucleotides; Protein Conformation; Pyrrolidines - chemistry; Pyrrolidines - pharmacokinetics; Pyrrolidines - pharmacology; SARS-CoV-2 - drug effects; SARS-CoV-2 - enzymology; Severe acute respiratory syndrome coronavirus 2; Structural biology; Thymine - chemistry; Thymine - pharmacokinetics; Thymine - pharmacology; Uracil; Uridine; Uridine - metabolism; Uridine monophosphate; Vanadate; Viral Nonstructural Proteins - antagonists & inhibitors; Viral Nonstructural Proteins - chemistry; Viral Nonstructural Proteins - metabolism
• ISSN: 2399-3642; 2399-3642
• DOI: 10.1038/s42003-021-01735-9

SARS-CoV-2 Nsp15 is a uridine-specific endoribonuclease with C-terminal catalytic domain belonging to the EndoU family that is highly conserved in coronaviruses. As endoribonuclease activity seems to be responsible for the interference with the innate immune response, Nsp15 emerges as an attractive target for therapeutic intervention. Here we report the first structures with bound nucleotides and show how the enzyme specifically recognizes uridine moiety. In addition to a uridine site we present evidence for a second base binding site that can accommodate any base. The structure with a transition state analog, uridine vanadate, confirms interactions key to catalytic mechanisms. In the presence of manganese ions, the enzyme cleaves unpaired RNAs. This acquired knowledge was instrumental in identifying Tipiracil, an FDA approved drug that is used in the treatment of colorectal cancer, as a potential anti-COVID-19 drug. Using crystallography, biochemical, and whole-cell assays, we demonstrate that Tipiracil inhibits SARS-CoV-2 Nsp15 by interacting with the uridine binding pocket in the enzyme’s active site. Our findings provide new insights for the development of uracil scaffold-based drugs. Youngchang Kim, Jacek Wower, and colleagues explore the sequence specificity, metal ion dependence and catalytic mechanism of the Nsp15 endoribonuclease NendoU from SARS-CoV-2. The authors also solve five new crystal structures of the enzyme in complex with 5’UMP, 3’UMP, 5’cGpU, uridine 2′,3′-vanadate (transition state analog) and Tipiracil (uracil mimic), and demonstrate that Tipiracil inhibits SARS-CoV-2 Nsp15 by interacting with the uridine binding pocket in the enzyme’s active site.