The amino acid sequence of vanillin-binding site of transient receptor potential vanilloid type 1 from rat, Leu 544 -Tyr 553 , was extracted and hybridized with His-tag. The hexadecamer invariant chain peptide, Leu-Ala-Met-Gly-Trp-Thr-Asn-Met-Leu-Tyr-His-His-His-His-His-His (VBH), was prepared by solid-phase peptide synthesis. Circular dichroic spectral measurements determined the α-helix content to be 17%, which was consistent to that of short peptides. In a combined use of thiol-derivatized nitrilotriacetic acid (s-NTA) monolayers, the His-tag successfully attached the whole peptide on gold substrate surfaces through Ni 2+ -chelation (Γ VBH = 224 ± 120 pmol cm -2 , n = 8). Moreover, various surface analyses including atomic force microscopy imaging, FT-IR spectroscopy, and quartz-crystal microgravimetry (QCM) revealed self-assembly (SA) of VBH at the S-NTA monolayer surfaces. QCM measurements also showed that vanillin, the major component of natural vanilla flavoring, binds to VBH SAs (K app = 2.7 × 10 3 M -1 ). The affinity of host-guest binding remains limited but possesses a certain degree of selectivity; for cases of structural analogs that give a pleasant flavor, acetophenone showed rather weak affinity (K app = 2.8 × 10 2 M -1 ) whereas 4-heptanone did not bind at all. With these results VBH was concluded to be useful in vanillin sensing as a supramolecular affinity host.

Herein, we report the self-assembly of a synthetic vanilloid receptor (VR) peptide that selectively binds capsaicin. We synthesized a 26-mer peptideYSEILFFVQS-HHHHHH-LAMGWTNMLY (S3HS4)comprising two chemoreceptor domains of transient receptor potential channel (TRPV1) linked by a hexahistidine sequence. High-speed atomic force microscopy (AFM) imaging in water revealed that the peptide structures alternated rapidly between wedge shape and linear forms. Circular dichroism spectroscopy showed that 65% of the amide units in the peptide chain adopted an α-helix structure, which was ascribed to the chemoreceptor domains. S3HS4 developed well-packed monolayers at the Ni-treated thiolated nitrilotriacetic acid self-assembled monolayers by chelation of the hexahistidine segment, as characterized by infrared spectroscopy and AFM, which exhibited statistically constant specific height. Therefore, S3HS4 was expected to fold spontaneously upon chelation, and the resulting helix–turn–helix conformers developed films while uniformly oriented: the tilt angle was 69° from the surface normal to the substrate. According to microgravimetric analysis using a quartz crystal microbalance (QCM), the adsorption was 84 ± 47 pmol cm–2 (n = 3), which was almost consistent with the saturation adsorption of an α-helix unit. We also used a QCM to investigate the host–guest reactions of S3HS4 and found that the S3HS4-attached QCM-chip-bound capsaicin with an apparent binding constant of (4.2 ± 3.6) × 104 M–1 (n = 4), whereas there was no evidence of binding to vanillin or acetophenone. Two controlsa blank chip without S3HS4 and a chip modified with a single helical peptide (LAMGWTNMLY-HHHHHH)produced no capsaicin response. To the best of our knowledge, S3HS4 is the first example of a synthetic VR mimic peptide. We believe that the present surface-directed structure-based design can be used to exploit the α-helix bundle in hexahistidine-linked bishelical peptides.