A Small Molecule, Lys-Ala-7-amido-4-methylcoumarin, Facilitates RNA Dimer Maturation of a Stem-Loop 1 Transcript in Vitro : Structure-Activity Relationship of the Activator

The type 1 human immunodeficiency virus (HIV-1), like all retroviruses, contains two copies of the RNA genome as a dimer. A dimer initially forms via a self-complementary sequence in the dimer initiation site (DIS) of the genomic RNA, but that dimer is converted to a mature dimer in a process generally promoted by the viral nucleocapsid (NC) protein. Formation of the mature dimer is correlated with infectivity. Study of genomic dimerization has been facilitated by discovery of short RNA transcripts containing the DIS stem-loop 1 (SL1), which can dimerize spontaneously without any protein factors in vitro as well as via the NC protein. On the basis of the palindromic nature of the apical loop of SL1, a kissing loop model has been proposed. First, a metastable kissing dimer is formed via a loop-loop interaction and then converted into a more stable extended dimer by the NC protein. This dimerization process in vitro is believed to mimic the in vivo RNA maturation. During experimental screening of potential inhibitors, we discovered a small molecule, Lys-Ala-7-amido-4-methylcoumarin (KA-AMC), which facilitates the in vitro conversion from kissing dimer to extended dimer. Here we report the structure-activity relationship for KA-AMC for promoting dimer maturation. Guanidino groups and increasing positive charge on the side chain enhance activity. For activity, the charged side chain is preferred on the benzene ring, and O 1 in the coumarin scaffold is essential. NMR studies show that the coumarin derivatives stack with aromatic bases of the RNA. The coumarin derivatives may aid in the investigation of some aspects of dimer maturation and serve as a scaffold for design of maturation inhibitors or of activators of premature maturation, either of which can lead to a potential HIV therapeutic.