Sulfer hot springs are a common source of archaebacteria, the third domain of li

Question

Sulfer hot springs are a common source of archaebacteria, the third domain of life that biohemically may to be more similar to eukaryotes than bacteria. You have just read research indicating that archaebacteria may be a rich source of a novel family of proteins, the Querall family. These proteins bind to selenium and activate other proteins through an unusual sulfhydryl modification of amines within the cell to repair telomeres, restore replicative ability, and as a result, reverse the aging process. Though they bind selenium, they seem to have a basic pI, in the range of 8.7 - 9.5. They also tend to be in the size range of the most common archae proteins, ranging from 10 kDa to 90 kDa. Eager to begin your study, you’ve already set up your own aritificial hot spring and have a thriving culture of archaebacteria to sample. However, you now need to isolate the proteins. a. Describe a method for isolating these proteins. Focus on the overall strategies and steps rather than specific buffers and lysis techniques, but be sure to be specific with your rationale for each technique. (15 points) b. What conditions you would use to test the enzyme activity? (5 points) c. You’re initial purified enzyme preparation appears to have little to no activity, though you’ve verified by tandem MS that your protein is present. What additional factors (other than the reaction conditions listed previously) might you need to consider? (5 points) Bonus points will be given if you incorporate a way to measure the enzyme activity into this answer.

Explanation / Answer

a) Lyse the cell and centrifuge at specific rpm to get the protein pellet. Then use the pellet to isolate the specific protein by Ion exchange chromatography using selenium. Use the pH less than their pI to make the protein positively charged

In cation exchange chromatography use selenium, an anion (Negatively charged) as resin (stationary phase) and the desired or specific protein (positively charged) will bind to the selenium.

b) pH equal to their pI and extreme temperature of above 1000 C

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