High Affinity Anti-inorganic Material Antibody Generation by Integrating Graft and Evolution Technologies

Abstract

Recent advances in molecular evolution technology enabled us to identify peptides and antibodies with affinity for inorganic materials. In the field of nanotechnology, the use of the functional peptides and antibodies should aid the construction of interface molecules designed to spontaneously link different nanomaterials; however, few material-binding antibodies, which have much higher affinity than short peptides, have been identified. Here, we generated high affinity antibodies from material-binding peptides by integrating peptide-grafting and phage-display techniques. A material-binding peptide sequence was first grafted into an appropriate loop of the complementarity determining region (CDR) of a camel-type single variable antibody fragment to create a low affinity material-binding antibody. Application of a combinatorial library approach to another CDR loop in the low affinity antibody then clearly and steadily promoted affinity for a specific material surface. Thermodynamic analysis demonstrated that the enthalpy synergistic effect from grafted and selected CDR loops drastically increased the affinity for material surface, indicating the potential of antibody scaffold for creating high affinity small interface units. We show the availability of the construction of antibodies by integrating graft and evolution technology for various inorganic materials and the potential of high affinity material-binding antibodies in biointerface applications.Peptides and proteins recognize the interfacial surfaces of their corresponding molecules with high affinity and selectivity because of the multiple-point interactions of hydrogen bonds and salt bridges and the surficial complementarities at the interfaces. Surface recognition by proteins has also been observed in biopolymers in biological systems (1, 2). Furthermore, the use of recent combinatorial library approaches has enabled the identification of short peptides with affinity for nonbiological inorganic materials (3-5). Peptides that bind materials such as metals, metal oxides, and semiconductors have been identified, and they are expected to be useful in bottom-up fabrication procedures in the field of bio-nanotechnology, such as patterning and assembly of proteins and nanomaterials (6 -8), biofunctionalization of nanoparticles (9, 10), and synthesis of crystalline nanometer-sized metal particles (11,12).Besides short peptides, antibodies are becoming attractive as novel material-binding molecules because they have higher affinities and specificities than peptides. Antibodies are recognition molecules with high binding affinity and specificity in the immune system, and they have been used widely in the fields of medical and analytical chemistry (13). By the use of general methodologies with in vivo immune system and in vitro combinatorial selection technologies, antibodies to the surfaces of organic crystals of 1,4-dinitrobenzene (14) and tripeptide (15), magnetite (16), gallium arsenide (17), gold (18), and polyhydroxybutyrate (19) have been identified in immuni...

Structural Guided Scaffold Phage Display Libraries as a Source of Bio-Therapeutics

“…More recently, a peptide sequence that bound to an inorganic material surface, was grafted into the CDR of a camel-type single domain antibody rearranged with a library of random sequences in additional CDR. Authors noted a synergistic effect from the grafted and selected random CDR loops that drastically increased the affinity for the inorganic target [23]. In this study we have taken a different approach, namely, grafting a 3-dimensional geometry based library designed from a ligand∶receptor binding stereochemical interface.…”

A High-Affinity Gold-Binding Camel Antibody: Antibody Engineering for One-Pot Functionalization of Gold Nanoparticles as Biointerface Molecules

“…33 The amino acids of the GBP-grafted CDR1 and randomized CDR3 are numbered as 1-1 to 1-18 and 3-1 to 3-16, respectively. 20 Bioconjugate Chemistry 23, 1934−1944 200 mM NaCl) containing 40 μg of ZnO particles (∼100 nm; Hosokawa Micron Inc., Hirakata, Japan) was sonicated for 1 min. E32 VHH, 4F2 VHH, and 4F2-E32 dimers were added to the suspension at various concentrations, and then 20 nm gold nanoparticles were added at a final concentration of 0.2 nM.…”

Enhancement of Cellulolytic Enzyme Activity by Clustering Cellulose Binding Domains on Nanoscaffolds

“…A CBD reversibly or irreversibly binds onto a substrate surface, and the binding is specific for crystalline or amorphous substrates. The thermodynamic factors associated with the binding of CBD onto cellulose depend on the surface of substrates: enthalpy changes drive binding onto an amorphous surface,27 such as in the cases of peptides and antibodies with affinities for inorganic material surfaces,28–30 whereas CBD binding to crystalline structures is entropy‐driven 31…”