Thermodynamics of three-way multibranch loops in RNA

RNA multibranch loops (junctions) are loops from which three or more helice s exit. They are nearly ubiquitous in RNA secondary structures determined b y comparative sequence analysis. In this study, systems in which two strand s combine to farm three-way junctions were used to measure the stabilities of RNA multibranch loops by UV optical melting and isothermal titration cal orimetry (ITC). These data were used to calculate the free energy increment for initiation of a three-way junction on the basis of a nearest neighbor model for secondary structure stability. Imino proton NMR spectra were also measured for two systems and are consistent with the hypothesized helical structures. Incorporation of the experimental data into the mfold and RNAst ructure computer programs has contributed to an improvement in prediction o f RNA secondary structure from sequence.