Biology Forums - Study Force

Recombinant expression of many mammalian proteins requires production of the
protein at below physiological temperatures (i.e. below 37 °C). If the protein is
expressed at 37 °C, all you produce is a large mass of aggregated protein called an
inclusion body. If you express the same protein at 20 °C, you produce a significant
amount of soluble, functional protein.

a. Provide a thermodynamically based reason for why this would be true.


Some proteins cannot be produced at all in bacteria but can be produced effectively in
either yeast or insect cells.

b. What is different about these cell lines that would allow them to produce
mammalian proteins better than bacteria?


c. What does this tell you about the folding pathway of these proteins?

(a) Proteins produced at higher temperatures denature the proteins by disrupting bonds that hold the molecules together. At lower temperatures, the bonds are less susceptible to molecular phase changes or more strongly held together. Think of ice water versus liquid water - the molecules in ice water are more strongly held together, but as soon as you apply heat, it increases disorder, causing the molecules to spread.

(b) Mammals have organelles that prompt proper protein folding, such as the endoplasmic reticulum. Bacteria don't have this luxury.

(c) Folding pathways in eukaryotes (yeast) are more structured/advanced/complex since they are modulated by numerous interactions with a variety of factors, including proteins such as chaperonins. This gives rise to more thermodynamically stable end-products.