Antigen specific T-cell Peptide MHC complex T-cell receptor HIV specific

HIV derived peptide-MHC

T-cell tagging strategy pioneered by Altman et al Peptide MHC complex Multimeric Binding Complex Antigen specific T-cell

T-cell tagging strategy pioneered by Altman et al Antigen specific T-cell

CLAIM 12: said multimeric binding complex is formed by binding said monomer to a multivalent entity through specific attachment sites Proimmune Pro5 Pentamer Pentameric entity with 5 peptide-MHC complexes capable of binding to T-cell receptor molecules Peptide-MHC complex Multivalent coiled-coil domain Site where pMHC monomer is specifically bound to the coiled-coil domain

Non-polar surfaces The Pro5 coiled-coil domain is a multivalent entity “Multivalency describes the binding of two (or more) entities that involves the simultaneous interaction between multiple, complementary functionalities on these entities” Mulder et al, Org. and Biomol. Chem., (2004) What holds the coiled-coil domain together? Hydrogen bonds Hydrophobic interactions Acceptor Donor Multiple, simultaneous interactions between complementary functionalities hold the helices together

Multimerization methods in ‘363 specification (6:3-5) “Preferably, the multimer will be formed by binding the monomers to a multivalent entity through specific attachment sites” (6:6-7) “The multimer may also be formed by chemical cross-linking of the monomers” (6:20-23) “The attachment site for binding to a multivalent entity … will be a specific binding pair member or one that is modified to provide a specific binding pair member, where the complementary pair has a multiplicity of specific binding sites.” (6:23-25) “Binding to the complementary binding member can be a chemical reaction, epitope-receptor binding, or hapten-receptor binding. (6:58-59) “The multivalent binding partner may be free in solution, or may be attached to an insoluble support.”

Multivalency Self-multimerization binding epitope receptor specific binding pair Coiled-coil from Pro5 Pentamer Interactions span the length of the helices and exist between all members of the complex There is no “specific binding pair” There is no “multivalent binding partner”