Protiens

Lecture 6 Working with proteins OBJECTIVES: At the end of these lectures, you should be subject to: mark methods of separation and purification Describe mainstay chromatography Describe isoelectric focusing Explain quantification of proteins Describe analysis of proteins by SDS/ rogue Describe western blotting Why purify a protein? To claim its function To analyze its physical properties To determine its rate For industrial or therapeutic applications Protein Purification Object: to complexify a especial(a) protein from all some other proteins and cell components thither are many an(prenominal) proteins (over 4300 genes in E. coli) A given protein stooge be 0.001-20% of summarize protein Other components: nucleic acids, carbohydrates, lipids, little(a) molecules Proteins are found in different states and locations: alcohol-soluble, insoluble, tissue layer bound, DNA bound, in organelles, cytoplasmic, periplasmic, atomic Protein s differ in: Size Shape germinate Properties stairs IN THE PURIFICATION OF A TYPICAL oil-soluble PROTEIN 1. Homogenization: manipulate crude extract. 2. Differential Centrifugation: Remove membranes, nuclei, greathearted organelles (mitochondria), and so on carrel free extract 3- Fractionation establish on protein size, charge, etc.

Many factors play role: pH, temperature, salt conc., and other factors 4. ammonium sulfate precipitation: (1) Proteins often become little soluble when the ionic strength is increased to very postgraduate levels. fall points vary by protein, so purification can be achieved. (2) Precipitates can be re-dissolved in a teensy volume, so concen tration can be achieved. (3) Dialyze the re-! dissolved proteins against a mild salt buffer. - Dialysis is a procedure that separate proteins (large size) from small solutes 5. newspaper column Chromatography First you need protein from cells: Differential Centrifugation Protein purification: tower chromatography -The nearly powerful method for fractionation - Solid var. (Matrix): porous...If you want to come up a full essay, order it on our website: OrderEssay.net

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