Im planning on using Ion Exchange Chromagraphy by DEAE-cellulose column for purifying an unknown cellulase from soil bacteria. How do i decide on the pH of the column buffer if protein pI is unknown? Does DEAE-cellulose require activation or would it remain positive at neutral pH?
Your question would be answered by experimentation. So, set up the experiment with appropriate conditions and controls. I am sure you will find all the answers.
Cellulase is not an unknown enzyme but rather your source is from an unknown organism.
Normally you would get your idea on the conditions used for purifctn from published data from similar studies
e.g.
Chidi, S. B., Godana, B., Ncube, I., Van Rensburg, E. J., Cronshaw, A., & Abotsi, E. K. (2008). Production, purification and characterization of celullase-free xylanase from Aspergillus terreus UL 4209. African Journal of Biotechnology, 7(21).
However, you might need to use further purification tool (Gel filtration) to purify the enzyme. For beginners, I would suggest to read Robert K. Scopes book on enzyme purification to understand some of the basics.
My book" Basic Enyzme Purification With Work Examples"
https://www.upmpress.com.my/product/basic-enyzme-purification-with-work-examples/
can be a rough guide.
As an MSc student, I guess you aim for your thesis is partial purification, of which ion exchange via DEAE-cellulose is probably enough.
Please look at the following below links which may help you in your analysis:
https://core.ac.uk/download/pdf/191971651.pdf
https://www.ncbi.nlm.nih.gov/books/NBK22410/
https://jb.asm.org/content/jb/174/13/4350.full.pdf
https://www.sheffield.ac.uk/polopoly_fs/1.779623!/file/PurificationGuide.pdf
Thanks
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