I am doing this undergraduate project on isolating and purification of bromelain enzyme but I am having difficulty in the concentration of buffer to use, the amount of the sephacel, pH of the buffer, the concentration of the elution buffer. I think I have left some out but please do anything in your power to help me.
Please if you don't have for DEAE-sephacel, you can help me on DEAE-cellulose or DEAE-sepharose.
You first need to look for the isoelectric point of your enzyme and if its activity depends on or it is affected by certain ions.
Please the isoelectric point for my enzyme is 4.6 but I can't tell if I should prepare the tris buffer to a pH of 6.5 Omar Gonzalez-Ortega
Tris buffer should be prepared around pH 8. You can find a typical procedure here:
http://wolfson.huji.ac.il/purification/pdf/ionexchange/amersham_ion_exchangemanual.pdf
What about the ions required or that are inhibitory?
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