GST-fusion problems

Arnoud van Vliet avvliet at knoware.nl
Thu Mar 8 06:02:07 EST 2001


> Duncan, which protease inhibitor(s) would you recommend if it is
> proteolysis, AEBSF..?

Best advice I can give is not use GST. I used it a lot, and always had
problems with degradation and insolubility. The ad says 70% soluble, I'd say
a max of 50%. And those are usually the controls :)

I made 10 overlapping clones of one gene, and 9 out of 10 were insoluble and
gave a lot of degradation products especially on Western blots. It was the
inserted part which was less stable. Check the figures in: van Vliet et al
(1994) Infect Immun 62:1451-1456 and van Vliet et al (1995) J Clin Microbiol
33:2405-2410.

I switched to 6His vectors, and never regretted it! This is probably not
what you wanted to hear, sorry for that.

hope this helps
Arnoud







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