Hey Biochemists,

How to extract Superoxide Dismutase from E coli. Need help.

Any help will be appreciated

Do you need an experimental protocol or what? Can you be specific?

Sparrow

Sparrow,
Yes, I need the experimental protocol.
I did not specify because I thought any other information might also be helpful.

.
The gene for human superoxide dismutase (hSOD), a cytoplasmic enzyme, was cloned into a high expression, secretion vector, pIN−III−OmpA. Upon induction of the gene expression, a new protein was produced at a level of approximately 10% of total cellular protein, which migrated at the identical position in SDS−polyacrylamide gel electrophoresis as hSOD produced in the E. coli cytoplasm. The product was recovered as the major component in the periplasmic fraction. The specific enzymatic activity of the secreted SOD was approximately half that of the cytoplasmic SOD expression with a non−secretion vector. These results indicate that a substantial fraction of the secreted SOD was able to be properly folded into the enzymatically active conformation. This is the first report to demonstrate that a foreign cytoplasmic protein can be secreted across the inner membrane into the periplasmic space yielding an active enzyme.
.

This might be helpful. Not for sure.

http://iai.asm.org/cgi/reprint/62/2/657.pdf

Sparrow

are u sure there is a protocal for this extraction from E.coli,????
would the protocol for any other bacteria work???

these basically exist as isozymes

Thank you very much sparrow, sayami and kancha.

I will be given E. coli and am asked to extract and purify that enzyme. I have about 3 hrs/week for 8 weeks to complete all.

I am going to sleep now. I will check this back tomorrow. Thank you again.

Copycat

Do u do copy and paste all the time...and that's why your nick is copycat? ;)

WELL, To me you sound like an undergarduate student( biology related major). If you are not, please disregard my posting.

I am assuming this is an project given to you( I was given the similar project when I was an undergraduate long time back) and honestly you should be doing some literature review not browsing sajha every other minute to see if sombody replied your thread with an answer

Its a learning experiment dear, and as we have the very true saying"Ki parera janincha ki padera janincha" and specially when you are in the feild of Biochem/ biology major, this is the only way. There is no alternative.

My advice to you,
1.go to "http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?DB=pubmed"
2.Type in "superoxide dismutase"
3. there are hundreds of paper,narrow down the search
4. Just scan thro' the title of the papers
5. scan thro' the Abstract of the paper
BOOOM ....there is your answer.

Good luck :)

and you do need to go through the details in the paper under "Methods and materials" and follow the reference if all the details you need is not in there.

Just out of curiosity....do u go to one of the schools in MN?

Good luck

thanks canabis. I just found out that you have commented in every thread I've asked help about academics

Well...well I tried what I could potentially do to help someone out, and Yes I figured being nerd is not that bad, or is it? ;). Its fun to get into academics converstaions somtimes =).