Human DEAD-box ATPase DDX3 shows a relaxed nucleoside substrate specificity.
Human DDX3 (hDDX3) is a DEAD-box protein shown to possess RNA-unwinding and adenosine triphosphatase (ATPase) activities. The hDDX3 protein has been implicated in nuclear mRNA export, cell growth control, and cancer progression. In addition, a role of this protein in the replication of human immunodeficiency virus Type 1 and in the pathogenesis of hepatitis C virus has been recently proposed. Its enzymological properties, however, are largely unknown. In this work, we characterized its ATPase activity. We show that hDDX3 ATPase activity is stimulated by various ribo- and deoxynucleic acids. Comparative analysis with different nucleoside triphosphate analogs showed that the hDDX3 ATPase couples high catalytic efficiency to a rather relaxed substrate specificity, both in terms of base selection and sugar selection. In addition, its ability to recognize the L-stereoisomers of both 3' deoxy- and 2',3' dideoxy-ribose, points to a relaxed stereoselectivity. On the basis of these results, we hypothesize the presence of structural determinants on both the base and the sugar moieties, critical for nucleoside binding to the enzyme. Our results expand the knowledge about the DEAD-box RNA helicases in general and can be used for rational design of selective inhibitors of hDDX3, to be tested as potential antitumor and antiviral agents.
Franca
Raffaella
R
DNA Enzymology and Molecular Virology Unit, Istituto di Genetica Molecolare IGM-CNR, via Abbiategrasso 207, 27100 Pavia, Italy.
Belfiore
Amalia
A
Spadari
Silvio
S
Maga
Giovanni
G
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D053487:DEAD-box RNA Helicases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D009696:Nucleic Acids / Q000378:metabolism; D009705:Nucleosides / Q000378:metabolism*; D011685:Purine Nucleotides / Q000494:pharmacology; D011742:Pyrimidine Nucleotides / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012680:Sensitivity and Specificity; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D009696:Nucleic Acids; D009705:Nucleosides; D011685:Purine Nucleotides; D011742:Pyrimidine Nucleotides; D011994:Recombinant Proteins; D008345:Manganese; D000251:Adenosine Triphosphatases; C119332:DDX3X protein, human; D053487:DEAD-box RNA Helicases; D008274:Magnesium