Genetic Characterization and Role in Virulence of the Ribonucleotide Reductases of Streptococcus sanguinis

Abstract Streptococcus sanguinis is a cause of infective endocarditis and has been shown to require a Mn transporter called SsaB for virulence and O2 tolerance. Like certain other pathogens, S. sanguinis possesses aerobic class Ib (NrdEF) and anaerobic class III (NrdDG) ribonucleotide reductases (RNRs) that perform the essential function of reducing ribonucleotides to deoxyribonucleotides. The accompanying paper indicates that in the presence of O2, the S. sanguinis class Ib RNR self-assembles an essential diferric-tyrosyl radical (FeIII2-Y*) in vitro whereas assembly of a dimanganese-tyrosyl radical (MnIII2-Y*) cofactor requires NrdI, and that MnIII2-Y* is more active than FeIII2-Y* with the endogenous reducing system of NrdH and thioredoxin reductase (TrxR1). In this study, we have shown that deletion of either nrdHEKF or nrdI completely abolishes virulence in an animal model of endocarditis, whereas nrdD mutation has no effect. The nrdHEKF, nrdI, and trxR1 mutants fail to grow aerobically whereas anaerobic growth requires nrdD. The nrdJ gene encoding an O2-independent, adenosylcobalamin-cofactored RNR was introduced into the nrdHEKF, nrdI, and trxR1 mutants. Growth of the nrdHEKF and nrdI mutants in the presence of O2 was partially restored. The combined results suggest that MnIII2-Y*-cofactored NrdF is required for growth under aerobic conditions and in animals. This could explain in part why Mn is necessary for virulence and O2 tolerance in many bacterial pathogens possessing a class Ib RNR, and suggests NrdF and NrdI may serve as promising new antimicrobial targets.