Length dependence of RNA-RNA annealing

The association of complementary nucleic acids can be described by a second order rate constant k. For extended molecules, including complex nucleic a cids, values of k were shown to be proportional to the square root of the c hain length L of the shorter nucleic acid strand at temperatures between t( m) and t(m) - 30 degrees C. For homopolymers this is true over a wider temp erature range. Below temperatures of t(m) - 30 degrees C, annealing rate co nstants may sharply decrease due to the formation of intramolecular structu res. It seems to be reasonable to assume that the formation of intramolecul ar structures of nucleic acids reduces the density of nucleation sites for annealing and, thereby, lowers the rates of association. Here, we examined the relationship between RNA chain length and the kinetics of RNA-RNA annea ling at physiological ionic strength and temperature. We used a complete se quence space derived from chloramphenicol acetyltransferase (cat) sequences to average over all structures for each given length. For groups of progre ssively longer antisense RNA species and a 800 nucleotides long complementa ry RNA, the observed annealing rate constants k(obs) were measured in vitro . The structure-averaged values for k(obs) of RNA-RNA annealing were not re lated to the square root of the chain length. Instead, they were found to b e proportional to 10(alpha L) (alpha = 0.0017). Here, a theoretical model i s suggested in which the observed length dependence is mainly influenced by ionic interactions between complementary RNA strands. The observed length dependence has substantial implications for the biological behavior of long -chain complementary RNA including the design of antisense RNA. The efficac y of antisense RNA in living cells is known to be related to annealing kine tics in vitro. Thus, on a statistical basis and independent of individual s tructures, long-chain rather than short-chain antisense RNA should lead to stronger inhibition. (C) 1999 Academic Press.