I'm going to cloning and overexpress a 7.5 kDa protein in E. coli base on pET vector system. But the problem is the low molecular weight proteins rapidly degraded by E. coli proteases. What should I do?
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How to obtain high-level production of low molecular weight recombinant proteins
Started by le_duy211, Dec 14 2011 12:39 AM
1 reply to this topic
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Posted 15 December 2011 - 10:55 PM
Hola, first made the production assay and if it fails think in change your sequence to a vector with periplasmic signal as pET 12, 20, 22 ,25 , 26, 27 and more advanced and look for a compatible protease deficient strain but BL21 is deficient in lon and omp proteases;after harvesting recover your protein of the periplasmic space by osmotic shock.Other possibility is fuse yor sequence to Gst and after purification digest the fusion protein. Buena suerte
Edited by protolder, 15 December 2011 - 10:56 PM.
