Mizuki Takahashi

Peptide design based on an antibody complementarity-determining region (CDR): construction of porphyrin-binding peptides and their affinity maturation by a combinatorial method.

We have utilized sequence information from an antiheme monoclonal antibody to develop novel porphyrin-binding peptides. Several peptides which have an intramolecular disulfide bond in different positions and different chain lengths were prepared. The affinities of peptides for meso-tetrakis(4-carboxyphenyl)porphyrin were increased by an appropriate conformational restraint using a disulfide bond. Detailed studies with a representative 12-peptide, 12C4, whose length was reduced from 20 residues of the complementarity-determining region (CDR), indicated that both the hydrophobic and electrostatic interactions were essential factors in the peptide-porphyrin binding. Moreover, two-dimensional 1H NMR spectroscopy revealed the conformation of the peptide and the critical residues for the porphyrin-binding. According to the obtained results, a further minimized 9-peptide, 9L, was successfully redesigned with a sequence capable of forming a beta-turn instead of a disulfide bond. Furthermore, affinity maturation studies of 9L were performed by using a combinatorial approach such as the spot-synthesis method. Peptides with an improved affinity for porphyrins were prepared by systematic amino acid replacement. Thus, the design of peptides targeted to porphyrins was demonstrated by the combination of antibody information and the rationally designed combinatorial method.

Design of novel porphyrin-binding peptides based on antibody CDR

Novel porphyrin-binding peptides were designed on the basis of an antigen binding site of an antiheme monoclonal antibody. Synthetic peptides were modified with a pyrene moiety. The spectroscopic measurements revealed that the synthetic peptides bound a porphyrin effectively.

Design of peptides derived from anti-IgE antibody for allergic treatment.

We have designed and synthesized peptides derived from an anti-IgE antibody which has a potential for the treatment of allergy. It was indicated that conformational restriction of peptide via an intramolecular disulfide bond improved the binding affinity for IgE and that the peptide might have an ability to inhibit the IgE-receptor interaction.

Construction of the novel conformationally-restricted peptide library for screening of peptides that control the interaction between nucleobases.

A unique conformationally-restricted peptide library was constructed using a loop structure as a structural scaffold. This library was used for the screening of the amino acid sequences that control the interaction between nucleobase triplets. The peptides have PNAs at the C-terminus as the recognition site and the random amino acid sequence at the N-terminus as the effector for the interaction between PNA and its complementary DNA triplets. From the peptide libraries constructed by the positional scanning method, the sequences that affect the interaction between PNA and complementary DNA were selected. The difference in the characteristic results by using A-T and G-C pairs was presented. This study would also give us some useful information about interaction between peptides and nucleic acids, such as relevances between these biomolecules in a prebiotic era.

Screening of Peptides that Control Interaction Between Nucleobases from Peptide Libraries Based on Loop Structures

The combinatorial peptide library has been widely used as a useful tool for screening of the bioactive ligands and/or recognition motif against specific molecules [1]. The random peptide libraries, however, generally have considerable conformational flexibility, thus many kinds of conformationally restrained peptide libraries have been reported recently [2]. In this study, we constructed a unique confomationally-restricted peptide library using a loop structure as a structural scaffold and peptide nucleic acids (PNAs) as a recognition site. By using this structure, we carried out the effective screening of the amino acid sequences that control the interaction between nucleobases.

Construction of porphyrin-binding peptides mimicking anti-heme monoclonal antibody

An antigen binding site of antibody (complementarity determining region; CDR) might have some structural or sequential factors to bind antigen. It has been reported that synthetic peptides derived from CDRs sequences have binding properties similar to the intact antibody [1, 2]. On the basis of CDR sequence and structure of monoclonal antibody against heme (FeIII-protoporphyrin IX), we attempted to construct new type of porphyrin-binding peptides. The interaction between porphyrins and synthetic peptides was studied by absorption, fluorescence and circular dichroism (CD) spectroscopies.