ADP-ribosylation of guanosine by SCO5461 protein secreted fromStreptomyces coelicolor

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Abstract

The Streptomyces coelicolor A3(2) genome encodes a possible secretion protein, SCO5461, that shares a 30% homology with the activity domains of two toxic ADP-ribosyltransferases, pierisins and mosquitocidal toxin. We found ADP-ribosylating activity for the SCO5461 protein product through its co-incubation with guanosine and NAD+, which resulted in the formation of N2-(ADP-ribos-1-yl)-guanosine (ar2Guo), with a Km value of 110 μM. SCO5461 was further found to ADP-ribosylate deoxyguanosine, GMP, dGMP, GTP, dGTP, and cyclic GMP with kcat values of 150–370 s−1. Oligo(dG), oligo(G), and yeast tRNA were also ADP-ribosylated by this protein, although with much lower kcat values of 0.2 s−1 or less. SCO5461 showed maximum ADP-ribosylation activity towards guanosine at 30 °C, and maintained 20% of these maximum activity levels even at 0 °C. This is the first report of the ADP-ribosylation of guanosine and guanine mononucleotides among the family members of various ADP-ribosylating enzymes. We additionally observed secretion of the putative gene product, SCO5461, in liquid cultures of S. coelicolor. We thus designated the SCO5461 protein product as S. coelicolor ADP-ribosylating protein, ScARP. Our current results could offer new insights into not only the ADP-ribosylation of small molecules but also signal transduction events via enzymatic nucleoside modification by toxin-related enzymes.

Highlights

★ ADP-ribosylating toxins mainly target proteins. However, pierisins ADP-ribosylates guanine residues in dsDNA. ★ SCO5461 protein, which shares homology with activity domains of MTX and pierisins, targets guanosine and deoxyguanosine. ★ We discovered SCO5461 for the first time as an enzyme which modulates nucleosides. ★ We discovered a possibly extracellular function of the ADP-ribosylation reaction.

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