4QFV

Crystal structure of a unique ankyrin

Summary for 4QFV

• Entry DOI: 10.2210/pdb4qfv/pdb
• Related: 4O60
• Descriptor: ANK-N5C-281, ACETATE ION (3 entities in total)
• Functional Keywords: designed ankyrin repeats, de novo protein, domain swap
• Biological source: synthetic construct
• Total number of polymer chains: 4
• Total formula weight: 100622.84

Authors

Ethayathulla, A.S.,Tikhonova, E.B.,Guan, L. (deposition date: 2014-05-21, release date: 2015-05-06, Last modification date: 2023-09-20)

Primary citation

Tikhonova, E.B.,Ethayathulla, A.S.,Su, Y.,Hariharan, P.,Xie, S.,Guan, L.
A transcription blocker isolated from a designed repeat protein combinatorial library by in vivo functional screen.
Sci Rep, 5:8070-8070, 2015

PubMed Abstract: A highly diverse DNA library coding for ankyrin seven-repeat proteins (ANK-N5C) was designed and constructed by a PCR-based combinatorial assembly strategy. A bacterial melibiose fermentation assay was adapted for in vivo functional screen. We isolated a transcription blocker that completely inhibits the melibiose-dependent expression of α-galactosidase (MelA) and melibiose permease (MelB) of Escherichia coli by specifically preventing activation of the melAB operon. High-resolution crystal structural determination reveals that the designed ANK-N5C protein has a typical ankyrin fold, and the specific transcription blocker, ANK-N5C-281, forms a domain-swapped dimer. Functional tests suggest that the activity of MelR, a DNA-binding transcription activator and a member of AraC family of transcription factors, is inhibited by ANK-N5C-281 protein. All ANK-N5C proteins are expected to have a concave binding area with negative surface potential, suggesting that the designed ANK-N5C library proteins may facilitate the discovery of binders recognizing structural motifs with positive surface potential, like in DNA-binding proteins. Overall, our results show that the established library is a useful tool for the discovery of novel bioactive reagents.

PubMed: 25627011
DOI: 10.1038/srep08070

Experimental method

X-RAY DIFFRACTION (1.999 Å)