incaRNAfbinv: a web server for the fragment-based design of RNA sequences

Abstract

In recent years, new methods for computational RNA design have been developed and applied to various problems in synthetic biology and nanotechnology. Lately, there is considerable interest in incorporating essential biological information when solving the inverse RNA folding problem. Correspondingly, RNAfbinv aims at including biologically meaningful constraints and is the only program to-date that performs a fragment-based design of RNA sequences. In doing so it allows the design of sequences that do not necessarily exactly fold into the target, as long as the overall coarse-grained tree graph shape is preserved. Augmented by the weighted sampling algorithm of incaRNAtion, our web server called incaRNAfbinv implements the method devised in RNAfbinv and offers an interactive environment for the inverse folding of RNA using a fragment-based design approach. It takes as input: a target RNA secondary structure; optional sequence and motif constraints; optional target minimum free energy, neutrality and GC content. In addition to the design of synthetic regulatory sequences, it can be used as a pre-processing step for the detection of novel natural occurring RNAs. The two complementary methodologies RNAfbinv and incaRNAtion are merged together and fully implemented in our web server incaRNAfbinv, available at http://www.cs.bgu.ac.il/incaRNAfbinv.

Figure 1.

Input screen of the incaRNAfbinv web server, configured for the design of a guanine-binding riboswitch aptamer (5), used as a pre-processing step in a novel riboswitch detection procedure (13). In addition to an input structure and sequence constraints, optional parameters include: motif selection for the fragment-based design, target minimum free energy, target mutational robustness, generation method for the seed (incaRNAtion is the default), GC content, number of simulated annealing iterations and number of output sequences.

Figure 2.

The results screen of the incaRNAfbinv web server, where the designed sequences are found in a table with options to sort and filter by selected parameters. Each row provides a designed sequence result and its folded predicted structure in dot-bracket notation (12), its Shapiro tree-graph representation (33), minimum free energy in kcal/mol, mutational robustness, base pair distance from input structure, Shapiro distance from input structure and an option to view the secondary structure drawing of its folded predicted structure using VARNA (36).

Figure 3.

Runtimes for four example structures: (i) miRNA-146 precursor (65 bases). (ii) Guanine-binding riboswitch aptamer (69 bases). (iii) Cobalamin riboswitch (127 bases). (iv) S14 Ribosomal RNA—Domain 2 (361 bases, for timing purposes). Each of the following structures was tested using five different GC% contents. The number of sequences designed was 20 by default. Tests were made using the default parameters and are presented in Log-10 s.

Figure 4.

Comparison of the free-energies of candidate solutions along the execution of RNAfbinv, for targeted GC contents from 0.1 to 0.9 and using incaRNAtion (solid lines) and the default random initialization of RNAfbinv (dashed lines), for the design of a guanine-binding riboswitch aptamer. Values averaged over 1000 runs.