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How to obtain high-level production of low molecular weight recombinant proteins - (Dec/14/2011 )

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?


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