Affinity Capture-Western

An interaction is inferred when a bait protein is affinity captured from cell extracts by either polyclonal antibody or epitope tag and the associated interaction partner identified by Western blot with a specific polyclonal antibody or second epitope tag. This category is also used if an interacting protein is visualized directly by dye stain or radioactivity. Note that this differs from any co-purification experiment involving affinity capture in that the co-purification experiment involves at least one extra purification step to get rid of potential contaminating proteins.

Publication

MCM double hexamer loading visualized with human proteins.

Weissmann F, Greiwe JF, Puehringer T, Eastwood EL, Couves EC, Miller TCR, Diffley JFX, Costa A

Eukaryotic DNA replication begins with the loading of the MCM replicative DNA helicase as a head-to-head double hexamer at origins of DNA replication1-3. Our current understanding of how the double hexamer is assembled by the origin recognition complex (ORC), CDC6 and CDT1 comes mostly from budding yeast. Here we characterize human double hexamer (hDH) loading using biochemical reconstitution and cryo-electron ... [more]

Nature Dec. 01, 2024; 636(8042);499-508 [Pubmed: 39604733]

Throughput

  • Low Throughput

Related interactions

InteractionExperimental Evidence CodeDatasetThroughputScoreCurated ByNotes
ORC1 ORC3
Affinity Capture-MS
Affinity Capture-MS

An interaction is inferred when a bait protein is affinity captured from cell extracts by either polyclonal antibody or epitope tag and the associated interaction partner is identified by mass spectrometric methods.

High-BioGRID
-
ORC1 ORC3
Affinity Capture-MS
Affinity Capture-MS

An interaction is inferred when a bait protein is affinity captured from cell extracts by either polyclonal antibody or epitope tag and the associated interaction partner is identified by mass spectrometric methods.

High-BioGRID
1452335
ORC3 ORC1
Reconstituted Complex
Reconstituted Complex

An interaction is inferred between proteins in vitro. This can include proteins in recombinant form or proteins isolated directly from cells with recombinant or purified bait. For example, GST pull-down assays where a GST-tagged protein is first isolated and then used to fish interactors from cell lysates are considered reconstituted complexes (e.g. PUBMED: 14657240, Fig. 4A or PUBMED: 14761940, Fig. 5). This can also include gel-shifts, surface plasmon resonance, isothermal titration calorimetry (ITC) and bio-layer interferometry (BLI) experiments. The bait-hit directionality may not be clear for 2 interacting proteins. In these cases the directionality is up to the discretion of the curator.

High-BioGRID
-

Curated By

  • BioGRID