Bio-Rad bulletin 5646A, 'An unrivaled, novel tag system for the expression of fusion-tagged proteins', describes the new cloning and expression kits for the Bio-Rad Profinity Exact fusion-tag system
Profinity Exact cloning and expression kits are part of the Profinity Exact fusion-tag system designed to address bottlenecks in procedures for purification and tag-removal of fusion-tagged proteins.
The bulletin describes how the Profinity Exact pPal7 expression vectors facilitate the expression of a unique N-terminal tag sequence encoding a built-in cleavage recognition site, occurring at the C-terminus of the tag.
The N-terminal Profinity Exact affinity tag is the prodomain of the Subtilisin protease.
Both tag and protease have been extensively modified for optimal purification results and exhibit a high degree of affinity (KD< 100pm).
The Subtilisin protease has been cleverly immobilised onto the purification resin for binding of Profinity Exact fusion-tagged proteins.
Cleavage by Subtilisin is triggered by fluoride buffers, occurs on-column and directly after the cleavage recognition site, and leaves a native protein behind.
With cleavage incubation times as short as 30 minutes, the Profinity eXact system provides a novel alternative to purify and process fusion proteins without the addition of protease.
Bulletin 5646A is available from either a local Bio-Rad sales office or it may be downloaded from the Bio-Rad website.
