A New Type of YumC-Like Ferredoxin (Flavodoxin) Reductase Is Involved in Ribonucleotide Reduction.
The trxB2 gene, which is annotated as a thioredoxin reductase, was found to be essential for growth of Lactococcus lactis in the presence of oxygen. The corresponding protein (TrxB2) showed a high similarity with Bacillus subtilis YumC (E value = 4.0E-88), and YumC was able to fully complement the ΔtrxB2 mutant phenotype. YumC represents a novel type of ferredoxin (flavodoxin) reductase (FdR) with hitherto-unknown biological function. We adaptively evolved the ΔtrxB2 mutant under aerobic conditions to find suppressor mutations that could help elucidate the involvement of TrxB2 in aerobic growth. Genome sequencing of two independent isolates, which were able to grow as well as the wild-type strain under aerated conditions, revealed the importance of mutations in nrdI, encoding a flavodoxin involved in aerobic ribonucleotide reduction. We suggest a role for TrxB2 in nucleotide metabolism, where the flavodoxin (NrdI) serves as its redox partner, and we support this hypothesis by showing the beneficial effect of deoxynucleosides on aerobic growth of the ΔtrxB2 mutant. Finally, we demonstrate, by heterologous expression, that the TrxB2 protein functionally can substitute for YumC in B. subtilis but that the addition of deoxynucleosides cannot compensate for the lethal phenotype displayed by the B. subtilis yumC knockout mutant.


OBJECTIVE
Ferredoxin (flavodoxin) reductase (FdR) is involved in many important reactions in both eukaryotes and prokaryotes, such as photosynthesis, nitrate reduction, etc. The recently identified bacterial YumC-type FdR belongs to a novel type, the biological function of which still remains elusive. We found that the YumC-like FdR (TrxB2) is essential for aerobic growth of Lactococcus lactis. We suggest that the YumC-type FdR is involved in the ribonucleotide reduction by the class Ib ribonucleotide reductase, which represents the workhorse for the bioconversion of nucleotides to deoxynucleotides in many prokaryotes and eukaryotic pathogens under aerobic conditions. As the partner of the flavodoxin (NrdI), the key FdR is missing in the current model describing the class Ib system in Escherichia coli. With this study, we have established a role for this novel type of FdR and in addition found the missing link needed to explain how ribonucleotide reduction is carried out under aerobic conditions.
National Food Institute, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark junch@food.dtu.dk chso@food.dtu.dk.
Jun
J
Chen
National Food Institute, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
Jing
J
Shen
National Food Institute, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark junch@food.dtu.dk chso@food.dtu.dk.
Christian
C
Solem
National Food Institute, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
Peter Ruhdal
PR
Jensen
D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D019020:Directed Molecular Evolution; D004926:Escherichia coli / Q000235:genetics; D005288:Ferredoxins / Q000378:metabolism; D005418:Flavodoxin / Q000378:metabolism; D016680:Genome, Bacterial; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D009154:Mutation; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D012264:Ribonucleotide Reductases / Q000235:genetics / Q000378:metabolism*; D012265:Ribonucleotides / Q000378:metabolism*; D016415:Sequence Alignment; D017421:Sequence Analysis
D001426:Bacterial Proteins; D005288:Ferredoxins; D005418:Flavodoxin; D012265:Ribonucleotides; D012264:Ribonucleotide Reductases; D010100:Oxygen