A human antibody fragment with high affinity for biodegradable polymer film.
Antibodies with high affinity for the surface of a solid material would be advantageous in biomaterial science as a protein device. A human antibody fragment that binds to poly(hydroxybutyrate) (PHB), a biodegradable polymer matter, was generated by a phage display system. Clone PH7-3d3 was isolated after several rounds of selection and prepared as a fragment of immunoglobulin variable regions (Fv). The quartz crystal microbalance technique showed that PH7-3d3 Fv completely inhibited PHB enzymatic degradation by competing with PHB depolymerase. Kinetic analysis based on surface plasmon resonance demonstrated that PH7-3d3 Fv bound to the PHB film with an equilibrium dissociation constant of 14 nM. The three-dimensional structure of PH7-3d3 Fv was resolved to 1.7 A, revealing that the complementarity determining regions (CDRs) in the Fv fragment form a relatively flat surface on which uncharged polar and aromatic amino acids are distributed in clusters. The structure of PH7-3d3 Fv was similar to that of PHB depolymerase in the orientation of aromatic residues in the binding sites. Alanine scanning mutagenesis demonstrated that these aromatic residues, especially tryptophan residues in CDRs, were critical in the interaction between PH7-3d3 Fv and PHB. Our results suggest the possible selection of an antibody fragment that binds a material surface in a manner similar to protein-ligand interaction.
Watanabe
Hideki
H
Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Aoba-yama 6-6-11-606, Sendai 980-8579, Japan.
Tsumoto
Kouhei
K
Taguchi
Seiichi
S
Yamashita
Koichi
K
Doi
Yoshiharu
Y
Nishimiya
Yoshiyuki
Y
Kondo
Hidemasa
H
Umetsu
Mitsuo
M
Kumagai
Izumi
I
D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D001673:Biodegradation, Environmental; D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors / Q000276:immunology*; D018360:Crystallography, X-Ray; D006801:Humans; D006885:Hydroxybutyrates / Q000737:chemistry*; D007128:Immunoglobulin Fragments / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D007135:Immunoglobulin Variable Region / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D019151:Peptide Library; D011091:Polyesters / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry; D000091402:Prohibitins; D011487:Protein Conformation; D020349:Surface Plasmon Resonance; D013499:Surface Properties; D000094122:Polyhydroxybutyrates
D000906:Antibodies; D002265:Carboxylic Ester Hydrolases; D006885:Hydroxybutyrates; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D011091:Polyesters; D011108:Polymers; D000091402:Prohibitins; C000717328:PHB protein, human; C003182:poly-beta-hydroxybutyrate; D000094122:Polyhydroxybutyrates; C022417:poly-beta-hydroxybutyrate depolymerase