Fluorescent indomethacin-dansyl conjugates utilize the membrane-binding domain of cyclooxygenase-2 to block the opening to the active site.

Publication Type:

Journal Article


J Biol Chem (2019)


<p>Many indomethacin amides and esters are cyclooxygenase-2- (COX-2)-selective inhibitors, providing a framework for the design of COX-2-targeted imaging and cancer chemotherapeutic agents. Although previous studies have suggested that the amide or ester moiety of these inhibitors binds in the lobby region, a spacious alcove within the enzyme&#39;s membrane-binding domain, structural details have been lacking. Here, we present observations on the crystal complexes of COX-2 with two indomethacin-dansyl conjugates (compounds 1and 2) at 2.22 Å resolution. Both compounds are COX-2-selective inhibitors with ICvalues of 0.76 μM and 0.17 μM, respectively. Our results confirmed that the dansyl moiety is localized in and establishes hydrophobic interactions and several hydrogen bonds with the lobby of the membrane binding domain. We noted that in both crystal structures, the linker tethering indomethacin to the dansyl moiety passes through the constriction at the mouth of the COX-2 active site, resulting in displacement and disorder of Arg-120, located at the opening to the active site. Both compounds exhibited higher inhibitory potency against a COX-2 R120A variant than against the wild-type enzyme. Inhibition kinetics of compound 2were similar to those of the indomethacin parent compound against wild-type COX-2, and the R120A substitution reduced the time dependency of COX inhibition. These results provide a structural basis for the further design and optimization of conjugated COX reagents for imaging of malignant or inflammatory tissues containing high COX-2 levels.</p>

6BL4 and 6BL3