Can a human recombinant glycoprotein produced in mammalian cell culture lose part of its sialic acid residues due to changing conditions in the culture or the purification of the protein? Any reading, institution or author you would recommend. Thanking you in advance
Sialic acids can be released from glycoproteins by acid hydrolysis at high temperature. So, I would try to avoid keeping cells in the same medium for more than 3 days and use a more basic buffer for purification steps. Here is a nice study - http://www.jbc.org/content/277/20/17502.full
Which cell line and which cell culture medium are you using? Can you share your purification protocol?
Thank you very much for your insightful response, Kavita. The cell line is CHO. I don't know which medium is used because I am troubleshooting the analysis of the purified glycoproteins to exclude degradation during analysis. But I want to give some recommendations to the cell culture scientists. Regarding purification, I know the harsher conditions used include steps using 2-propanol or ammonia at 2-8 °C or 0.5M ammonium hydrogen carbonate buffer. Do you think any of those may induce sialic acid release from glycoproteins? When you said high temperature, how many degrees celsius did you mean? Thanks again for sharing your knowledge.
Sorry for the late response. Not sure if you have figured out the problem already.
' Do you think any of those may induce sialic acid release from glycoproteins? ' - I am not sure. May be you can use a purified glycoprotein like Fetuin (available commercially) - process it through same conditions and see if that leads to loss of sialic acid or degradation.
'When you said high temperature, how many degrees celsius did you mean?'- I use 80 degrees for 3 hrs.
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