Abstract
The helicase from severe acute respiratory syndrome coronavirus (SARS-CoV) possesses NTPase, duplex RNA/DNA-unwinding and RNA-capping activities that are essential for viral replication and proliferation. Here, we have isolated DNA aptamers against the SARS-CoV helicase from a combinatorial DNA library. These aptamers show two distinct classes of secondary structure, G-quadruplex and non-G-quadruplex, as shown by circular dichroism and gel electrophoresis. All of the aptamers that were selected stimulated ATPase activity of the SARS-CoV helicase with low-nanomolar apparent K(m) values. Intriguingly, only the non-G-quadruplex aptamers showed specific inhibition of helicase activities, whereas the G-quadruplex aptamers did not inhibit helicase activities. The non-G-quadruplex aptamer with the strongest inhibitory potency was modified at the 3'-end with biotin or inverted thymidine, and the modification increased its stability in serum, particularly for the inverted thymidine modification. Structural diversity in selection coupled to post-selection stabilisation has provided new insights into the aptamers that were selected for a helicase target. These aptamers are being further developed to inhibit SARS-CoV replication.