Difference between revisions of "Part:BBa K782007"

Revision as of 13:54, 25 September 2012

NicTAL12:NLS DNA binding domain

Introduction

TAL effectors (TALEs) are bacterial plant pathogen transcription factors, that bind to DNA by specifically recognizing one base pair with a single tandem repeat in their DNA-binding domain. A tandem TALE repeat contains 33 to 35 amino acids, where the 12th and 13th amino acid, called a “repeat variable diresidue” (RVD), are responsible for specific interactions with the corresponding base pair (Scholze and Boch, 2011).

Figure 1: Schematic representation of the construct

Characterization

We discovered that NicTAL part deposited in the Registry by the Slovenian iGEM2010 team [1] was missing subdomain in DNA-binding domain, so our team added this missing part on N terminal sequence. Apart from adding N terminal sequence,we added HIS tag on N terminal end and NLS on terminal C end of NicTAL12 (Figure 2).

This construct was later used for designing TAL-based activator and repressor by adding VP16 [1] and KRAB [2] domain.

Results obtained by testing NicTAL12 repressor and activator, proved that NicTAL12 binds to its bonding sites (link do BS) better than NicTAL10.

Figure 3: results of testing NicTAL12:VP16 activator and NicTAL12:KRAB repressor, shows, that NicTAL12 binds to DNA better than NicTAL10.

Single binding sequence for NicTAL:

References

Scholze, H., and Boch, J. (2011) TAL effectors are remote controls for gene activation. Curr. Opin. Microbiol. 14, 47-53.

Sequence and Features