The p-cumaric acid esterase (p-CAE) has a wide range of applications and is therefore an enzyme of interest. p-CAE was produced by Pichia pastoris and purified by immobilized metal ion affinity chromatography (IMAC). Different IMAC systems were tested and compared. 35.4% of the total protein was identified as active p-CAE. A total activity loss of 21.4% was recorded. The p-CAE could be successfully purified.
Elution chromatograms at 280 nm of four IMAC systems and SDS PAGE of eluate (1) and eluate treated with Endo H (2)
Validation of four IMAC systems by recovery rate for enzyme activity and protein content
Optimal IMAC system with IDA and pH 8 for purification of p-CAE; A) Chromatogramm 280nm, B) SDS page (M - marker, F - filtrate, BT - breakthrough, W - wash, E - eluat, EEH - eluat treated with Endo H)
| Method | FPLC |
|---|
| Mode | Affinity |
|---|---|
| Substances | p-cumaric acid esterase |
| Key words | FPLC, Affinity, AZURA FPLC, IMAC, protein purifiaction |
| CAS number | - |
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