Bke2 Biochemistry Lectures

Lecture 8: Biochemical methods
Reading material: Horton, Chapter 3, and protein purification lab manual

Abstract: Chromatography and electrophoresis are commonly used methods for purification and analysis of proteins. In gel-filtration chromatography, large molecules go faster through the column than small ones, so that molecules of different size may be separated. With SDS-polyacrylamide gel electrophoresis small molecules move faster than larger. Ion exchange chromatography separates molecules according to their charge. Molecular mass can be determined by ultracentrifugation, gel filtration and most accurately by electrospray mass spectrometry. Protein sequences can be determined. Electron microscopy is a valuable method to show the structure of cell organelles and larger macromolecules. X-ray crystallography is the outstanding method to determine detailed molecular structures from small molecules to very large viruses. NMR can be used to determine smaller macromolecular structures in solution.

Key concepts:
Electrophoresis (native or denaturing gels)
Dialysis
Ammonium sulfate precipitation
Chromatography
Ion exchange chromatography
Gel filtration chromatography
Affinity chromatography
Centrifugation
Spectroscopic methods

Lecture index

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