BCH210H1 Lecture 17: L17
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BCH210H1 Full Course Notes
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Refolding of ribonuclease: after dialysis to remove small molecules (urea and mercaptoethanol), the enzyme slowly regained activity, denatured proteins can refold to regain their native structure, the sulfhydryl groups became re-oxidized by air to form disulfide bonds. Dialysis: contains a semipermeable membrane, small molecules (like; urea, mercaptoethanol) have the ability to pass through the membrane, protein subunits are too large to pass through, therefore they will remain on one side. In order to renature the protein: undergo dialysis to remove urea and. Mercaptoethanol: air oxidation to allow the reformation of disulfide bonds. Christian anfinsen: experiment conclusion: results with ribonuclease led to the main conclusion that, amino acid sequence specifies final protein conformation. Primary structure determines secondary and tertiary structure: when rnase was refolded in 8m urea by oxidation, and then dialyzed to remove urea only 1% enzymatic activity was obtained. This was due to the wrong disulfide pairing. There are 105 ways to pair 8 cysteines into 4 disulfide pairs.
