Scabin, a novel DNA-acting ADP-ribosyltransferase from Streptomyces scabies

July 28, 2017


Scabin, a novel DNA-acting ADP-ribosyltransferase from Streptomyces scabies


Bronwyn Lyons, Ravikiran Ravulapalli, Jason Lanoue, Miguel R. Lugo, Debajyoti Dutta, Stephanie Carlin, A. Rod Merrill




The Journal of Biological Chemistry


A bioinformatics strategy was used to identify Scabin, a novel DNA-targeting enzyme from the plant pathogen 87.22 strain of Streptomyces scabies. Scabin shares nearly 40% sequence identity with the Pierisin family of mono-ADP-ribosyltransferase toxins. Scabin was purified to homogeneity as a 22-kDa single-domain enzyme and was shown to possess high NAD+-glycohydrolase (Km(NAD) = 68 ± 3 μM; kcat = 94 ± 2 min−1) activity with an RSQXE motif; it was also shown to target deoxyguanosine and showed sigmoidal enzyme kinetics (K0.5(deoxyguanosine) = 302 ± 12 μM; kcat = 14 min−1). Mass spectrometry analysis revealed that Scabin labels the exocyclic amino group on guanine bases in either single-stranded or double-stranded DNA. Several small molecule inhibitors were identified, and the most potent compounds were found to inhibit the enzyme activity with Ki values ranging from 3 to 24 μM. PJ34, a well known inhibitor of poly-ADP-ribosyltransferases, was shown to be the most potent inhibitor of Scabin. Scabin was crystallized, representing the first structure of a DNA-targeting mono-ADP-ribosyltransferase enzyme; the structures of the apo-form (1.45 Å) and with two inhibitors (P6-E, 1.4 Å; PJ34, 1.6 Å) were solved. These x-ray structures are also the first high resolution structures of the Pierisin subgroup of the mono-ADP-ribosyltransferase toxin family. A model of Scabin with its DNA substrate is also proposed.




Circular dichroism, Secondary structure, Protein folding, Biochemistry