Abstract
© Springer Science+Business Media New York 2015. All rights are reserved. All researchers immersed in the world of recombinant protein production are in agreement that often the production and purification process of a protein can become a nightmare due to an unexpected behavior of the protein at different protocol stages. Once the protein is purified, scientists know that they still cannot relax. There is a decisive last step missing: performing a protein dialysis in a suitable buffer for subsequent experimental trials. Here is when we can find proteins that precipitate during dialysis by buffer-related factors (ionic strength, pH, etc.), which are intrinsic to each protein and are difficult to predict. How can we find the buffer in which a protein is more stable and with less tendency to precipitate? In this chapter we go over possible factors affecting the protein precipitation tendency during the dialysis process and describe a general dialysis protocol with tricks to reduce protein aggregation. Furthermore, we propose a fast method to detect the most appropriate buffer for the stability of a particular protein, performing microdialysis on a battery of different buffers to measure afterwards precipitation by a colorimetric method, and thus being able to choose the most suitable buffer for the dialysis of a given protein.
Original language | English |
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Title of host publication | Insoluble Proteins: Methods and Protocols |
Pages | 321-330 |
Number of pages | 9 |
DOIs | |
Publication status | Published - 1 Dec 2014 |
Keywords
- Dialysis
- Microdialysis
- Protein aggregation
- Protein precipitation
- Protein stability