title: Self-assembly of protein monolayers engineered for improved monoclonal immunoglobulin G binding.
abstract: Bacterial outer membrane proteins, along with a filling lipid molecule can be modified to form stable self-assembled monolayers on gold. The transmembrane domain of Escherichia coli outer membrane protein A has been engineered to create a scaffold protein to which functional motifs can be fused. In earlier work we described the assembly and structure of an antibody-binding array where the Z domain of Staphylococcus aureus protein A was fused to the scaffold protein. Whilst the binding of rabbit polyclonal immunoglobulin G (IgG) to the array is very strong, mouse monoclonal IgG dissociates from the array easily. This is a problem since many immunodiagnostic tests rely upon the use of mouse monoclonal antibodies. Here we describe a strategy to develop an antibody-binding array that will bind mouse monoclonal IgG with lowered dissociation from the array. A novel protein consisting of the scaffold protein fused to two pairs of Z domains separated by a long flexible linker was manufactured. Using surface plasmon resonance the self-assembly of the new protein on gold and the improved binding of mouse monoclonal IgG were demonstrated.
authors.affiliation: Institute for Cell and Molecular Biosciences, The Medical School, Newcastle University, Framlington Place, Newcastle upon Tyne, NE2 4HH, UK; E-Mail:  abn@ansto.gov.au.
authors.forename: Anton P
authors.initials: AP
authors.lastname: Le Brun
authors.forename: Deepan S H
authors.initials: DS
authors.lastname: Shah
authors.forename: Dale
authors.initials: D
authors.lastname: Athey
authors.forename: Stephen A
authors.initials: SA
authors.lastname: Holt
authors.forename: Jeremy H
authors.initials: JH
authors.lastname: Lakey
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002942:Circular Dichroism; D003001:Cloning, Molecular; D007074:Immunoglobulin G / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding / Q000276:immunology; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D007074:Immunoglobulin G; D011506:Proteins; D011994:Recombinant Proteins; C078167:OMPA outer membrane proteins