Abstract: The present invention provides methods for crystallographic structure determination employing hydrogen exchange analysis. Hydrogen exchange analysis is used to identify unstructured regions of a protein, and then hydrogen exchange analysis repeated after the protein is admixed with candidate agents and/or conditions that may induce structure in said identified unstructured regions. Agents that induce desired structure in the protein are then employed, admixed with the protein, in co-crystallization studies for structure determination. Hydrogen exchange analysis is performed by determining the quantity of isotope and/or rate of exchange of peptide amide hydrogen(s) with isotope on a labeled protein. Proteins with agent-induced decreases in unstructured regions, and thus improved hydrogen exchange structural maps, are optimal for high quality crystallization and structure determination.

Abstract: The present invention provides methods for crystallographic structure determination employing hydrogen exchange analysis. Hydrogen exchange analysis is used to identify unstructured regions of a protein, which are then deleted to obtain one or more modified proteins for crystallization and structure determination. Optionally, an initial hydrogen exchange stability map of a protein is compared to that of at least one modified form of the protein to identify unstructured regions, while retaining characteristic structure of the native protein. Hydrogen exchange analysis is performed by determining the quantity of isotope and/or rate of exchange of peptide amide hydrogen(s) with isotope on a labeled protein. Proteins with fewer unstructured regions, and thus improved hydrogen exchange structural maps, are optimal for high quality crystallization and structure determination.

Abstract: The present invention provides methods whereby the positions of peptide amide groups that are labeled with a heavy hydrogen in a polypeptide or protein can be localized at high resolution. The methods are useful for determining which peptide amide groups in a polypeptide or protein are accessible to solvent, mapping the binding site and/or binding surface of a binding protein, and/or studying allosteric or other conformational changes in a polypeptide or protein which alter the rates at which certain peptide amide hydrogens exchange with solvent.

Abstract: The present invention provides methods of identifying hot-spot residues for one or both members of a receptor-ligand complex of interest. Further provided are methods of using receptor hot-spot residues to identify compounds that functionally bind a receptor in a manner that mimics the binding of a known ligand for the receptor.

Abstract: The binding sites of binding proteins and their binding partners are characterized, at the individual amino acid level, by a combination of tritium exchange labeling and sequential degradation and analysis of tritiated fragments under slowed exchange conditions.

Abstract: The present invention provides methods whereby the positions of peptide amide groups that are labeled with a heavy hydrogen in a polypeptide or protein can be localized at high resolution. The methods are useful for determining which peptide amide groups in a polypeptide or protein are accessible to solvent, mapping the binding site and/or binding surface of a binding protein, and/or studying allosteric or other conformational changes in a polypeptide or protein which alter the rates at which certain peptide amide hydrogens exchange with solvent.

Abstract: The binding sites of binding proteins and their binding partners are characterized, at the individual amino acid level, by a combination of tritium exchange labeling and sequential degradation and analysis of tritiated fragments under slowed exchange conditions.