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bioRxiv · 10.1101/373563

Fusion of a functional glutaredoxin to the radical-generating subunit of ribonucleotide reductase

Rozman Grinberg, I.·Lundin, D.·Sahlin, M.·Crona, M.·Berggren, G.·Hofer, A.·Sjöberg, B.-M.
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Abstract

Class I ribonucleotide reductase (RNR) consists of a catalytic subunit (NrdA) and a radical-generating subunit (NrdB) that together catalyse reduction of the four ribonucleotides to their corresponding deoxyribonucleotides. Facklamia ignava NrdB is an unprecedented fusion protein with N-terminal add-ons of a glutaredoxin (Grx) domain followed by an ATP-cone. Grx, which in general is encoded elsewhere in the genome than is the RNR operon, is a known physiological reductant of RNRs. Here we show that the fused Grx domain functions as an efficient reductant of the F. ignava class I RNR via the common dithiol mechanism and interestingly also via a monothiol mechanism, although less efficiently. A Grx that utilizes either or of these two reaction mechanisms has to our knowledge not been observed with a native substrate before. The ATP-cone, which is commonly found as an N-terminal domain of the catalytic subunit of RNRs, is an allosteric on/off switch that promotes dNDP reduction in presence of ATP and inhibits the enzyme activity in presence of dATP. Here we show that dATP bound to the ATP-cone of F. ignava NrdB promotes formation of tetramers that are unable to form enzymatically competent complexes with F. ignava NrdA. The ATP-cone binds two molecules of dATP, but only one molecule of the activating nucleotide ATP. F. ignava NrdB contains the recently identified radical factor Mn2III/IV. We show that NrdA from the firmicute F. ignava can form a catalytically competent RNR with the Mn2III/IV-containing NrdB from the flavobacterium Leeuwenhoekiella blandensis.

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Rozman Grinberg, I., Lundin, D., Sahlin, M., Crona, M., Berggren, G., Hofer, A., Sjöberg, B.-M.. 2018-07-20. Fusion of a functional glutaredoxin to the radical-generating subunit of ribonucleotide reductase. https://doi.org/10.1101/373563

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