The NIH guidelines for work with large volumes (> 10 liter) of genetically engin

Question

The NIH guidelines for work with large volumes (> 10 liter) of genetically engineered microorganisms requires that the organisms be killed or contained prior to further processing for product recovery. Fortunately, much of the recombinant DNA work is done in E. coli that is very sensitive to thermal death. A typical value for the activation energy for thermal death of E. coli is 75 kcal/mol. Many of the products of interest from genetically engineered E. coli are proteins: the activation energy for thermal denaturation of protein is typically 25 kcal/mol. The greater sensitivity of E. coli over proteins to thermal destruction suggests that heat treatment of E. coli at the end of the fermentation may provide a means of killing cells without causing substantial damage to the protein product. You may use a continuous sterilizer with steam injection to heat kill the cells to 10^-1 viable organisms/fermentor as they are pumped from the fermentor to a collection vessel. The 10 m^3 fermentor contains E. coli at 5 times 10^9 cell/ml and is operated at 37 degree C. If the sterilizer is operated at 62 degree C, what fraction of the protein in the cell will be denatured? Some useful data: Death constant for E. coli at 54 degree C K= 0.25 min^-1 Denaturation constant for protein at 40 degree C K_p = 5 times 10^-5 sec^-1 R = universal gas constant = 1.99 cal/mol degree K

Explanation / Answer

Activation energy for thermal death of E.coli=75kcal/mol

Activation energy for thermal denaturation of protein=25kcal/mol

Death constant for E.coli K=0.25min-1

R=1.99cal/mol degree K

Denaturation constant for protein at 40degree kp=5*10^-5 sec-1

The fraction of protein denaturated=(activation energy for protein denaturation/activation energy for thermal death of E.coli)*(denaturation constant /death constant)

=(25/75)*(5*10^-5/0.25*60)*R

=0.004*1.99

=0.0079cal/mol degree K

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