Hello,
I tried different methods of making refolding buffer including cystine, I dissolve it in HCL first but then when I raise the pH to 8 I can see it solidifying after raising the pH. how can I handle cystine solution?
Dear Kimia Kaffashi ,
The problem is that this substance is intrinsically poorly soluble in water and therefor there is an upper limit which is ‘simply’ there. For the maximum you can do, you might have a look at: Solubilities of l-Cystine, l-Tyrosine, l-Leucine, and Glycine in Aqueous Solutions at Various pHs and NaCl Concentrations. Renzo Carta and Giuseppe Tola Journal of Chemical & Engineering Data 1996 41 (3), 414-417 DOI: 10.1021/je9501853 (see enclosed file).
Overall message: high salt will improve solubility but, in all cases, only in the extreme regions of pH (low or high), in other words once you are back in a buffer round let’s say pH 7 then you will get precipitates.
As indicated in your related question here on RG: https://www.researchgate.net/post/Refolding_buffer_contain_cystine_and_cysteine_protocol
If nothing seems to work, you might consider alternatives for the cystine.
Best regards.
For reoxidizing disulfide bonds during refolding a "redox buffer" composed of a mixture of reduced and oxidized glutathione is often used, typically 3 mM reduced and 1 mM oxidized. Glutathione does not have the solubility problem you encountered with cystine at those concentrations.
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