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how to make protein soluble? - protein solubility (Apr/06/2011 )

Hi Forumers :D

Here I come again, with another problem....please forgive me :(

I have this problem in expressing the protein in soluble form. I've played with the IPTG concentration, the temperature during induction and also I've tried using lower temperature which is 22oC when I start incubate it until reach OD600=0.6. Is there any way to solve this? Please..any suggestions and recommendations are appreciated :D
Attached Image

-rpjkmust916-

rpjkmust916 on Thu Apr 7 05:56:47 2011 said:


Hi Forumers :D

Here I come again, with another problem....please forgive me :(

I have this problem in expressing the protein in soluble form. I've played with the IPTG concentration, the temperature during induction and also I've tried using lower temperature which is 22oC when I start incubate it until reach OD600=0.6. Is there any way to solve this? Please..any suggestions and recommendations are appreciated :D

Hola, I suposse that these images are at +-28ēC in LB with 0.2mM IPTG and 3 hours of induction. I think that your protein is tagged ,isn´t it?.What strain do you use? Have any expression before induce? your protein is about 30-40KD. I would use any Origami or Rosetta gami strain for expression, and I would chech any of the internet tools to predice solubility introducing aminoacid sequence if the probability of insoluble form ia quite high, bad thing my friend. Buena suerte

-protolder-

protolder on Thu Apr 7 07:07:08 2011 said:


rpjkmust916 on Thu Apr 7 05:56:47 2011 said:


Hi Forumers :D

Here I come again, with another problem....please forgive me :(

I have this problem in expressing the protein in soluble form. I've played with the IPTG concentration, the temperature during induction and also I've tried using lower temperature which is 22oC when I start incubate it until reach OD600=0.6. Is there any way to solve this? Please..any suggestions and recommendations are appreciated :D

Hola, I suposse that these images are at +-28ºC in LB with 0.2mM IPTG and 3 hours of induction. I think that your protein is tagged ,isn´t it?.What strain do you use? Have any expression before induce? your protein is about 30-40KD. I would use any Origami or Rosetta gami strain for expression, and I would chech any of the internet tools to predice solubility introducing aminoacid sequence if the probability of insoluble form ia quite high, bad thing my friend. Buena suerte



Gracias Protolder :D

You're almost right!!! I wrote I've incubated it at 22oC, forgive me to give wrong information. Actually, I've incubate it at 27oC, with 0.1mM IPTG for 4 hrs. It is a tagged protein with Rosetta strain. Could you explain about the internet tools to predict the solubility. How I can use it?

Muchas gracias :D

-rpjkmust916-

Hola, Now i have changed protein expression and purification for animal pharmacology. These are the tools that I used:
http://mips.helmholtz-muenchen.de/proso/proso.seam.
biotech.ou.edu, from Oklahoma unv.
refold.med.monash.edu.au
!que tengas un buen día ! (the first is .de/proso/proso.seam.)

-protolder-

Muchas Gracias Protolder :D

-rpjkmust916-