Means and methods for breaking noncovalent binding interactions between molecules

Inventors

Ovaa, Huib • Schumacher, Antonius Nicolaas Maria

Assignees

Stichting Sanquin Bloedvoorziening • Het Nederlands Kanker Instituut

Abstract

The present invention concerns multimeric proteinaceous molecules comprising at least two members that bind each other via a region of noncovalent interaction, wherein a first of the members comprises a conditionally reactive group that, when activated, cleaves a covalent bond within the first member. Cleavage of the covalent bond results in a reduction in the binding strength with which the at least two members bind to each other via the region of noncovalent interaction. The reduction in the binding strength can result in the separation of the members under mild conditions.

Core Innovation

The invention concerns breaking noncovalent interactions in multimeric proteinaceous molecules by incorporating a conditionally reactive group into a member of the multimer, wherein activation cleaves an internal covalent bond. Activation reduces noncovalent binding strength by cleaving the temporary peptide into at least two peptides that each exhibit reduced binding affinity for the MHC molecule with respect to the uncleaved temporary peptide.

The described problem is that exchanging peptides on MHC multimers/tetramers and generating ligand-occupied MHC for T-cell detection and diagnostic screening can be difficult to perform without harsh conditions that disrupt the multimeric proteinaceous molecule or otherwise compromise desired binding. The disclosed solution uses an MHC molecule provided with a temporary peptide that binds to the peptide-binding groove, where the temporary peptide contains a reactive group that is either a light-sensitive group or a periodate-sensitive group.

Upon activation of the light-sensitive or periodate-sensitive reactive group, the temporary peptide is cleaved into at least two peptides that exhibit reduced binding affinity for the MHC molecule, thereby reducing noncovalent interaction strength and enabling dissociation. The document emphasizes MHC tetramers and MHC multimers used for peptide exchange and for making ligand-occupied MHC complexes, including applications such as T-cell detection and diagnostic screening.

Claims Coverage

The provided independent claim set contains 1 independent claim. The inventive coverage is centered on producing an MHC molecule using a temporary peptide that can be cleaved upon activation by either a light-sensitive group or a periodate-sensitive group, generating cleaved peptides with reduced binding affinity to the MHC.

Overall, the claim coverage is directed to MHC molecule production in which a temporary peptide is bound in the peptide-binding groove and is selectively activated to cleave into multiple peptides with reduced affinity, thereby enabling peptide release and ligand-occupied MHC formation.

Stated Advantages

Documented Applications