Capture proteasome activity assay (CAPA)
Fig. 1: Measurement of the activity of proteasome captured from 293 cells expressing either SP, IP, SIP or DIP. Captured proteasome are incubated in the presence of commercially available fluorogenic substrates to measure their chymotrypsin, caspase and trypsin-like activities, respectively (A). The chymotrypsin-like activities of each proteasome type was measured in the presence of increasing concentration of the b5i-specific proteasome inhibitor PR957. The b5i subunit is present in IP, SIP and DIP , but not in SP (B).
Measurement of proteasome subtypes activity
Four cell lines were developed, each expressing one specific subtype of proteasome:
These cell lines were characterized using proprietary antibodies developed against the native subunits of proteasome1 and will be provided as cell lysates.
The CAPA test consists in capturing a specific proteasome subtype from the cell lysate on antibody-coated plates and using classic fluorogenic substrates for direct proteasome activity read-out.
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