Structure-Based Models to Understand the Cipher that Governs Tale Protein Affinity with DNA.

Abstract

Recent use of TALEs, transcription activator like effectors as a novel DNA binding domain, has raised the interest in deciphering the mechanisms of DNA recognition by these multimeric TALE proteins. TALE proteins can be fused with the catalytic domain of FOKI nucleases to act as artificial restriction enzymes to create a targeted DNA double strand break in-vivo for genome editing. The TAL effectors are characterized by a central domain of 30 tandem repeats of 33 to 35 amino acid sequence motifs. The amino acid sequence of each repeat is largely invariant, with the exception of two adjacent amino acid at position 12th and 13th (the repeat variable diresidue or RVD). Repeats with different RVD recognize different DNA base pairs, and it is hypothesized that there is one-to-one correspondence between the RVDs in the repeat domain and the nucleotide in the target DNA sequence. But how this central domain contacts DNA and what determines the specificity has remained enigmatic so far. Here we use structure-based models to investigate the structure of TALENS and their interaction with DNA. Simulations with biophysical models for multimeric protein in complex with DNA are presently too computationally expensive to help, but structure-based simulations been used to study protein folding and protein-protein interaction mechanisms for over a decade. We performed homology modeling to construct a structure of single repeat and 1.5 repeat using the PthA (TPR domain) as a template protein, which predicts only alpha helices and turns for both the repeat domains. Strucuture based models are generated by inspecting the native contacts of DNA and individual repeat domain from previous experimental knowledge and can be easily adapted to different DNA sequences and multidomain of TALE, as long as the binding motives are conserved.