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Purification of Hsp104, a Protein Disaggregase

17.2K Views

07:17 min

September 30th, 2011

September 30th, 2011

17,224 Views

0:05

Title

1:07

Cell Harvest and Lysis

1:55

Ni-affinity Purification of Hsp104

3:16

FPLC Purification of Hsp104

5:14

Representative Results for Hsp104 Purification and Assessment of Disaggregation Activity

7:04

Conclusion

Here, we describe a protocol for the purification of highly active Hsp104, a hexameric AAA+ protein from yeast, which couples ATP hydrolysis to protein disaggregation. This scheme exploits a His6-tagged construct for affinity purification from E. coli followed by anion-exchange chromatography, His6-tag removal with TEV protease, and size-exclusion chromatography.

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