Arranging quaternary structure of peptides by cyclodextrin-guest inclusion complex: sequence-specific DNA binding by a peptide dimer with artificial dimerization module

Abstract

Arranging quaternary structures by dimerization of monomers is necessary for many sequence-specific DNA-binding proteins to become functional. Upon dimer formation, the DNA contacting regions of each monomer are positioned to a proper orientation that facilitates efficient sequence-specific recognition of DNA.1 Several laboratories have reported model studies that demonstrate the importance of such steric constraints by using covalently bonded dimeric peptides.* 12 However, natural proteins dimerize with noncovalent interactions, and the equilibrium governing the formation of dimers would be important for regulation.3 Recent findings on the transcriptional control mechanisms by heterodimer formation for the basic leucine-zipper protein and the basic helix-loop-helix protein families also indicate that the specificity in dimer formation regulates the sequencespecific DNA binding of such proteins.4 W e report here synthetic oligopeptides that bind to specific DNA sequences upon noncovalent dimer formation. A peptide modified with /3-cyclodextrin (/3-CD) and a peptide with an adamantyl group form a heterodimer mediated by formation of an inclusion complex between the /3-CD and the adamantyl group, and the heterodimer binds to a specific DNA sequence.A 23-residue peptide (G23) derived from the DNA contacting region of the transcriptional activator protein GCN4,5* one of the large family of DNA-binding proteins characterized by a basic leucine-zipper structural motif, was used as a sequencespecific DNA-binding domain. GCN4 binds to the specific DNA sequence upon formation of dimer, which is mediated by a parallelcoiled coil assembly of each leucine-zipper domain.5 Our design for effecting noncovalent dimer formation is a host-guest inclusion complex of /3-CD and its guest compound. Modification of the C-terminal cysteine residue of G23 with mono-6-deoxy-6-iodo-/3-cyclodextrin6 afforded G23-CD, while modification with /V-(bromoacetyl)-1 -adamantanemethylamine gave G23-AD (Figure l).7 It has been shown that 1-adamantaneacetic acid forms a 1:1 inclusion complex in water with /3-cyclodextrin with a dissociation constant of 5 X 10~5 M.8 We have tested whether• To whom correspondence should be addressed.

A Highly Sensitive 27 MHz Quartz-Crystal Microbalance as a Device for Kinetic Measurements of Molecular Recognition on DNA Strands

“…These molecular recognitions have been mainly studied by gel mobility shift assay. [1][2][3][4][5] This technique is widely used in molecular biology; however, it has some difficulties with regard to quantitatively detecting interactions. DNA or peptide is usually labeled either directly or indirectly with fluorescent or radioactive molecules, and it takes relatively a long time to analyze the results.…”

Stimuli-responsive selection of target DNA sequences by synthetic bZIP peptides

“…Particularly relevant are those based on dimeric bZIP basic regions, in which the natural C/C-terminal leucine zipper is replaced by artificial dimerizers [14][15][16][17] . Some of these designs have gone even further and incorporate switchable elements that allow conditional off/on DNA binding by the application of external stimuli such as light or metal ions [18][19][20][21][22] .…”

Sequence-selective DNA recognition and enhanced cellular up-take by peptide–steroid conjugates

“…Building on this successful idea of miniaturisation, Morii, Schepartz, Mascareñas Q3 and our group have enforced the proof-of-concept by using a variety of small dimerizing moieties. [7][8][9][10][11][12] Subsequently, we have shown that the attachment of the basic region peptides to a rigid scaffold, a derivative of deoxycholic acid in this case, also allows selective recognition of DNA. Indeed, our previous work on cMyc-Max b-HLH-ZIP and GCN4-bZIP proteins showed that this type of steroid-based constructs show potential for binding DNA.…”