Characterization of the Drosophila melanogaster Dis3 ribonuclease.
The Dis3 ribonuclease is a member of the hydrolytic RNR protein family. Although much progress has been made in understanding the structure, function, and enzymatic activities of prokaryotic RNR family members RNase II and RNase R, there are no activity studies of the metazoan ortholog, Dis3. Here, we characterize the activity of the Drosophila melanogaster Dis3 (dDis3) protein. We find that dDis3 is active in the presence of various monovalent and divalent cations, and requires divalent cations for activity. dDis3 hydrolyzes compositionally distinct RNA substrates, yet releases different products depending upon the substrate. Additionally, dDis3 remains active when lacking N-terminal domains, suggesting that an independent active site resides in the C-terminus of the protein. Finally, a study of dDis3 interactions with dRrp6 and core exosome subunits in extracts revealed sensitivity to higher divalent cation concentrations and detergent, suggesting the presence of both ionic and hydrophobic interactions in dDis3-exosome complexes. Our study thus broadens our mechanistic understanding of the general ribonuclease activity of Dis3 and RNR family members.
Department of Molecular Biology and Microbiology and Program in Cell Biology, Case Western Reserve University School of Medicine, Cleveland, OH 44106, USA.
Megan
M
Mamolen
Erik D
ED
Andrulis
D000818:Animals; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000201:enzymology*; D004795:Enzyme Stability; D063326:Exosome Multienzyme Ribonuclease Complex; D055354:Exosomes / Q000201:enzymology*; D017434:Protein Structure, Tertiary / Q000235:genetics; D012260:Ribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
C502521:Dis3 protein, Drosophila; D029721:Drosophila Proteins; C545895:Rrp6 protein, Drosophila; D063326:Exosome Multienzyme Ribonuclease Complex; D012260:Ribonucleases