Stability of the Tyrosyl Radical in the Ribonucleotide Reductase Beta Subunit of Arcobacter bivalviorum

Arcobacter spp., such as Arcobacter bivalviorum (A. bivalviorum), are free-living organisms found in diverse environments and associated with animals. They are considered emerging enteropathogens and potential zoonotic agents. Ribonucleotide reductase (RNR) is the key enzyme that is used to convert ribonucleotides into deoxyribonucleoside triphosphates (dNTPs). This process utilises radical-based chemistry and is crucial for DNA biosynthesis and repair. There are three RNR classes, with class I RNR the most studied, present in A. bivalviorum, eukaryotes, and many prokaryotes. Class I RNRs are further divided into three subclasses: Ia, Ib, and Ic. Class Ib RNRs use a dimanganese-oxo centre, unlike class Ia RNRs, which use a diiron-oxo centre. A. bivalviorum possesses a class Ia enzyme that requires a diferric tyrosyl radical cofactor located within its beta (β) subunit. Indeed, both the efficiency and fidelity of DNA synthesis are influenced by the stability of the tyrosyl radical (Y•) in the RNR, which is a critical aspect of its enzymatic function. This study investigates the stability of the Y-radical (Y•) site within the RNR β subunit of A. bivalviorum and the nature of the neighbouring amino acid residues. To achieve these goals, we developed a model of the RNR β subunit of A. bivalviorum, using the RNR β subunit of Aquifex aeolicus as a reference template (7aik.1. A PDB). The results provide some important details about the radical site and its surrounding residues, highlighting the influence of the protein structure on the stability of the radical. These findings may guide the development of novel inhibitors targeting this enzyme in A. bivalviorum.