Abstract
Transcription activator-like (TAL) effectors specifically bind to double stranded (ds) DNA through a central domain of tandem repeats. Each TAL effector (TALE) repeat comprises 33–35 amino acids and recognizes one specific DNA base through a highly variable residue at a fixed position in the repeat. Structural studies have revealed the molecular basis of DNA recognition by TALE repeats. Examination of the overall structure reveals that the basic building block of TALE protein, namely a helical hairpin, is one-helix shifted from the previously defined TALE motif. Here we wish to suggest a structure-based re-demarcation of the TALE repeat which starts with the residues that bind to the DNA backbone phosphate and concludes with the base-recognition hyper-variable residue. This new numbering system is consistent with the α-solenoid superfamily to which TALE belongs, and reflects the structural integrity of TAL effectors. In addition, it confers integral number of TALE repeats that matches the number of bound DNA bases. We then present fifteen crystal structures of engineered dHax3 variants in complex with target DNA molecules, which elucidate the structural basis for the recognition of bases adenine (A) and guanine (G) by reported or uncharacterized TALE codes. Finally, we analyzed the sequence-structure correlation of the amino acid residues within a TALE repeat. The structural analyses reported here may advance the mechanistic understanding of TALE proteins and facilitate the design of TALEN with improved affinity and specificity.Electronic supplementary materialThe online version of this article (doi:10.1007/s13238-014-0035-2) contains supplementary material, which is available to authorized users.
Highlights
We wish to suggest a structure-based re-demarcation of the TAL effector (TALE) repeat which starts with the residues that bind to the DNA backbone phosphate and concludes with the base-recognition hyper-variable residue
We present fifteen crystal structures of engineered dHax3 variants in complex with target DNA molecules, which elucidate the structural basis for the recognition of bases adenine (A) and guanine (G) by reported or uncharacterized TALE codes
Transcription activator-like (TAL) effectors belong to the αsolenoid superfamily which includes pentatricopeptide repeat proteins (PPR) (Yin et al, 2013), tetratricopeptide repeat proteins (TPR) (Das et al, 1998), and other proteins
Summary
Transcription activator-like effector nuclease (TALEN) is becoming an important tool for genome-editing in multiple species (Bogdanove and Voytas, 2011; Huang et al, 2011; Carlson et al, 2012; McMahon et al, 2012; Streubel et al, 2012; Beumer et al, 2013; Christian et al, 2013; Doyle et al, 2013; Heigwer et al, 2013; Kim et al, 2013; Panda et al, 2013). TALENs exploit the DNA-binding domain of TAL effectors, which usually comprise 1.5 to 33.5 tandem repeats (TALE repeats), for sequence specific and customizable DNA recognition (Swarup et al, 1992; Bonas et al, 1993; Bai et al, 2000; Gu et al, 2005; White and Yang, 2009; Boch and Bonas, 2010). The sequence-specific RVD codes were deciphered through both experimental and bioinformatic investigations (Boch et al, 2009; Moscou and Bogdanove, 2009; Streubel et al, 2012; Yin et al, 2012; Yang et al, 2014). The well-characterized RVD codes include NI (Asn and Ile) for adenine (A), HD (His and Asp) for cytosine (C), NG (Asn and Gly) for thymine (T) and methylated cytosine (mC), NH (Asn and His) and NK (Asn and Lys) for guanine (G), NN (Asn and Asn) for G/A, and NS (Asn and Ser) for all the four bases (Boch and Bonas, 2010; Bogdanove and Voytas, 2011; Streubel et al, 2012; Yang et al, 2014)
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