9NCS

RNase A in complex with Uridine Vanadate and decavanadates

Summary for 9NCS

• Entry DOI: 10.2210/pdb9ncs/pdb
• Descriptor: Ribonuclease pancreatic, DECAVANADATE, URIDINE, ... (5 entities in total)
• Functional Keywords: ribonuclease, rna, uridine vanadate, decavanadate, hydrolase
• Biological source: Bos taurus (domestic cattle)
• Total number of polymer chains: 2
• Total formula weight: 31557.39

Authors

Gutierrez, C.S.,Lim, D.C.,Silkenath, B.,Kojasoy, V.,Raines, R.T. (deposition date: 2025-02-17, release date: 2025-09-24)

Primary citation

Gutierrez, C.S.,Silkenath, B.,Kojasoy, V.,Pich, J.A.,Lim, D.C.,Raines, R.T.
Pseudouridine Residues as Substrates for Serum Ribonucleases.
Rna, 2025

PubMed Abstract: In clinical uses, RNA must maintain its integrity in serum that contains ribonucleases (RNases), especially RNase 1, which is a human homolog of RNase A. These omnipresent enzymes catalyze the cleavage of the P-O bond on the 3' side of pyrimidine residues. Pseudouridine (Ψ) is the most abundant modified nucleoside in natural RNA. The substitution of uridine (U) with Ψ or ‑methylpseudouridine (mΨ) reduces the immunogenicity of mRNA and increases ribosomal translation, and these modified nucleosides are key components of RNA-based vaccines. Here, we assessed the ability of RNase A and RNase 1 to catalyze the cleavage of the P-O bond on the 3' side of Ψ and mΨ. We find that these enzymes catalyze the cleavage of UpA up to 10‑fold more efficiently than the cleavage of ΨpA or mΨpA. X-ray crystallography of enzyme-bound nucleoside 2',3'‑cyclic vanadate complexes and molecular dynamics simulations of enzyme·dinucleotide complexes show that U, Ψ, and mΨ bind to RNase A and RNase 1 in a similar manner. Quantum chemistry calculations suggested that the higher reactivity of UpA is intrinsic, arising from an inductive effect that decreases the p of the 2'‑hydroxy group of U and enhances its nucleophilicity toward the P-O bond. Experimentally, we found that UpA does indeed undergo spontaneous hydrolysis faster than does mΨpA. Our findings inform the continuing development of RNA-based vaccines and therapeutic agents.

PubMed: 40835455
DOI: 10.1261/rna.080404.125

Experimental method

X-RAY DIFFRACTION (1.83 Å)