Abstract
A nickel-chelate affinity chromatography has been used to isolate fused proteins of Hsp70-E7 from yeast lysate at the initial stage of the work. The low purity of the monomeric Hsp70-E7 in the final fraction was associated with the occurrence of dimeric and tetrameric forms of the target protein and low-molecular impurities. A modification of the purification procces by the addition of reduced L-glutathione to inhibit the formation of unwanted disulfide bonds permitted to increase the purity of the target protein by reducing the content of the oligomerized products. Further changes in the purification protocol, namely the reduction of hydrophobic interactions by the introduction of glycerol, significantly reduced the amount of low-molecular impurities. For a finer purification of E7-HSP70, a two-stage process of anion-exchange chromatography that made it possible to effectively eliminate low-molecular impurities was used. The assessment of the final material by electrophoresis under non-reducing conditions showed that the purity of the monomeric forms of either of the proteins was higher than 95%.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.