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BISC 330 Spring 2011 Lecture 8

BISC 330 Spring 2011 Lecture 8 - BISC 330L Biochemistry...

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BISC 330L: Biochemistry Spring Semester 2011 USC Lecture 8: Friday, 28 January 2011 1. Begin Chapter 3 material… Biochemistry Toolbox Techniques in Protein Analysis 3 0 L
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Chapter 3 : Exploring Proteins and Proteomes • Biochemistry Toolbox: Techniques in Protein Purification – Gaining access to proteins –Purifying according to 1. Solubility, 2. Size, 3. Charge, and 4. Binding Affinity – Centrifugation –Gel electrophoresis
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So you want to study Protein X… •Dec ide on a protein of interest • Develop assay for Protein X Î Enzyme act. • Determine total [protein] in mixture •Spec i f
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Proteins come from……. . CELLS Where do we get proteins from? • Therefore, a protein must be separated from the other cellular components before its function and structure can be further studied.
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Homogenate/ Cell extract Sonication How do we extract proteins from cells? •In order to release the cell’s contents we must first break (lyse) the cell membrane. This process is called homogenization . Detergent lysis Mortar/Pestle Force Now what do we do?!!!
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Fig. 3.1 Example of cell fractionation by Differential Centrifugation. (Dounce homogenizer + minced tissue in cold buffer) spin
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Density gradient ( Zonal ) centrifugation separates matters according to density . •Often provides a better resolution of
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BISC 330 Spring 2011 Lecture 8 - BISC 330L Biochemistry...

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