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Photocontrol of DNA Binding Specificity of a Miniature Engrailed Homeodomain

Guerrero, Lucia, Smart, Oliver S., Woolley, G. Andrew and Allemann, Rudolf Konrad ORCID: 2005. Photocontrol of DNA Binding Specificity of a Miniature Engrailed Homeodomain. Journal of the American Chemical Society 127 (44) , pp. 15624-15629. 10.1021/ja0550428

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Control of DNA binding of HDH-3, a 18-residue polypeptide based on the recognition helix of the Q50K engrailed homeodomain, has been achieved. HDH-3 was linked to an azobenzene cross-linker through two cysteine residues in an i, i + 11 spacing. For the thermodynamically stable trans configuration of the cross-linker, the dark-adapted peptide (dad-HDH-3) adopted a mainly α-helical structure as judged by circular dichroism (CD) spectroscopy. After irradiation with light of 360 nm, the helical content of the peptide (irrad-HDH-3) was reduced significantly and the CD spectrum of the irradiated peptide resembled that of the largely unstructured, unalkylated peptide. Despite lacking helices-1 and -2 and the N-terminal arm of Q50K engrailed, dad-HDH-3 bound to its natural DNA target sequence TAATCC (QRE) with high affinity (KD = 7.5 ± 1.3 nM). The binding affinity for the mutant DNA sequence, TAATTA (ERE), was reduced significantly (KD = 140 ± 11 nM). Unlike irrad-HDH-3, which like the unalkylated parent peptide displayed only marginal DNA binding specificity, dad-HDH-3 specified base pairs 5 and 6 of QRE with an accuracy rivaling that of the intact wild-type Q50K engrailed homeodomain, making dad-HDH-3 the most specific designed DNA binding miniature homeodomain reported to date. Moreover, DNA binding affinity and specificity of HDH-3 could be controlled externally by irradiation with light.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Cardiff Catalysis Institute (CCI)
Subjects: Q Science > QD Chemistry
Publisher: American Chemical Society
ISSN: 0002-7863
Last Modified: 18 Oct 2022 13:19

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