Technology

Technology 2017-03-28T10:13:49+00:00

What is DEPC Labeling?

QuarryBio’s technology is based on the reactivity of the small molecule DEPC with side chain functional groups of 6 different amino acids and the n-terminal amine.

The extent of DEPC labeling with each amino acid is primarily driven by the position of the amino acid in the protein structure. Amino acids located on the surface of a protein are highly exposed to the surrounding solvent that contains DEPC, and react to a greater extent than amino acids buried within the protein structure.

Because changes in solvent accessibility can be detected through changes in the extent of DEPC labeling, this technology is an effective tool for epitope mapping and identifying changes in protein higher order structure (HOS).

Diagram: DEPC labeling

Using DEPC for Epitope Mapping

Diagram: Using DEPC for epitope mapping

When an antigen is bound to a mAb or Fab, residues at the binding will have significantly reduced solvent accessibility, which will decrease the extent of DEPC labeling. Comparing DEPC labeling for an antigen both in isolation vs. the antigen bound to a mAb/Fab enables identification of these residues with decreased labeling, allowing ready identification of epitopes and sites of allosteric impact.

Using DEPC to identify changes in protein structure

Diagram: Using DEPC for HOS labeling

When a protein is subject to stressed conditions that result in a change in the protein’s 3 dimensional structure, the solvent accessibility of amino acids in the impacted regions are impacted. Comparing DEPC labeling of a protein in the native state vs. a protein subjected to stressed conditions enables identification of residues with changes in DEPC labeling, allowing ready identification of regions of the protein structure that were impacted by the stress conditions.

Performing DEPC labeling experiments

Diagram: DEPC labeling process

To perform a DEPC labeling experiments, the protein sample is incubated in a DEPC solution and then quenched. Proteolysis prepares the sample for LCMS analysis, which is able to identify peptides containing DEPC labels. MSMS analysis enables the identification of specific amino acids in each peptide that contain DEPC labels. Performing this DEPC labeling experiment on different protein samples enables the direct comparison of DEPC labeling levels, and identification of residues where labeling levels change.

DEPC labeling can be used with most common aqueous buffer solutions, and can accommodate proteins of essentially any size.