CHEM237 Lecture 5: CHEM237-5-Ch5-Purification-annot.PDF
10 May 2016
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Topics: source of protein making a lysate ion exchange gel filtration affinity chromatography. Hic chromatography salting out techniques chromatographic techniques electrophoresis techniques (sds) page electrophoresis isoelectric focusing. A series of steps to purify a protein from a source. Source of protein? considerations: concentration of protein in the source ease of isolation cost. Use of molecular cloning techniques can increase the concentration of protein in the cell enzyme assay: determine the activity of the enzyme protein detection: determine absolute amount of protein in the sample fermentation techniques cell culture techniques animals, plants. Specific activity = activity of the enzyme in units (e. g. moles of product of the enzyme reaction) per mg of protein in the solution. As purity goes up, the specific activity goes up. The protein becomes more pure as the purification proceeds. Percent recovery = amount of activity recovered compared to the total amount of activity present initially.
