multiple identical genes in an organism - (May/01/2007 )
Hello!
I heard that we can put multiple identical genes in an organism to increase the yield of one particular protein.
I am wondering, why are then researchers and companies trying to improve the yield of one proiten, if you can just put 5 identical genes in an organism.
Some companies are selling products that improve the of an protein. Why not put an extra copy in the organism and it is cheaper?
Thanksfor explanation
Marko
In theory.
In a world were there are rainbows day after day.
But lets say the protein is toxic? Or misfolded in the bacteria?
Or becomes insoluble in the cell when present at high concentration? Forming nasty aggregates that are tough to resolubilise and may be trapped in a misfolded state.
More gene copies won't save the day.
Furthermore lets say, the gene expression level varies with the energetic / metabolism / carbon utilisation / carbon -nitrogen balance? Thus I might get lots of protein when the cells access to certain quantity of oxygen and carbon and energy. Vary any one of this and protein products fall off.. sometimes dramaticaly sometimes not.
Now to add more problem what happen when these conditions also affect the fold state of the desired protein (Remember a protein is just a polypeptide sting you get after translation, it needs folding (which requires cheperon) and it interacts other proteins in its environment when folding, and it need post-translational modifications eg addition of polysaccaride moeties, lipids, reduction, addition of metal centres etc. So, if I grow the cells this way, I get functional protein. Grow it another way and all the protein comes in a horrible insoluble pellet.
Don't forget cultivation temperature. That too influences the state of the protein, even its yield levels. (Mainly due to toxicity)
And finally, people need a lot of protein quickly and usually don't want to or able to process lots of dilute cultures (100s Litres). Growing cells to high density bring about many of the above problems.
Yes, all that you said is true.
But, I wont to know why people bother to improve protein yields when only one copy of a protein is inserted.
Why are they playing around with pH, T, etc... when they could just put an extra copy of the gene and have the same yield.
I am wondering about differences between optimization of protein expression and simply puting more copies of a gene?
Some companies sell products that enhance protein translation(maybe they insert (GAA)8 into UTR).
Why not just use aditional copy of a gene?
But, I wont to know why people bother to improve protein yields when only one copy of a protein is inserted.
Why are they playing around with pH, T, etc... when they could just put an extra copy of the gene and have the same yield.
I am wondering about differences between optimization of protein expression and simply puting more copies of a gene?
Some companies sell products that enhance protein translation(maybe they insert (GAA)8 into UTR).
Why not just use aditional copy of a gene?
Why optimise protein expresion by modification of UTR sequence? I believe that one is new to me.
Why bother, is the first thing that comes to mind. I don't know. If one were over expressing a protein wouldn't one use strong promoters/enhanser units and a generic terminator sequnences. Furthermore the UTR is for the most part useless if I grabed a human gene and tried to express it in E coli, or yeast or insects cells. And UTR usually contain control elements, and protein are regulated strongly. Not the best thing to add for over expression.... hmmm...
No clue here.
I can see codon optimisation for the gene. If a xeno-gene used a rare codon for the host organism,then no matter how many copies or how strongly expressed the gene, the pool of the rare tRNA-amino acid remains the same size and continues to limit overall expression.
I can see using better promoter-enhanser constructs. Promoter strenght can very by several order of a magnitude, especially since many of our common strong promoters are generic, works in any beast (organism) but not actually the strongest possible expression level attainable.