SDS-PAGE Sodium Dodecyl Sulfate - Polyacrylamid Gel Electrophoresis Timothy G. Standish, Ph. D.

Sodium Dodecyl Sulfate SDS is a common ingredient in detergents Other names for SDS include lauryl sulfate and sodium lauryl sulfate As a detergent SDS destroys protein secondary, tertiary and quaternary structure This makes proteins rod shaped SDS also sticks to proteins in a ratio of approximately 1.4 g of SDS for each gram of protein Negative charge on the sulfate groups of SDS mask any charge on the protein

SDS Sodium Dodecyl Sulfate C12H25NaO4S Non-polar Hydrophobic tail Polar Hydrophilic head H-C-C-C-C-C-C-C-C-C-C-C-C-O-S-O-Na+ H O Because it is amphipathic, SDS is a potent detergent SDS may also be called sodium lauryl sulfate and is a common ingredient of shampoos, laundry detergent and other cleaning products

SDS and Proteins SDS Protein

SDS and Proteins SDS nonpolar chains arrange themselves on proteins and destroy secondary tertiary and quarternary structrure Thus shape is no longer an issue as the protein SDS complex becomes rod shaped In aqueous solutions, SDS polarizes releasing Na+ and retaining a negative charge on the sulfate head So much SDS binds to proteins that the negative charge on the SDS drowns out any net charge on protein side chains In the presence of SDS all proteins have uniform shape and charge per unit length

Polyacrylamide Gels Polyacrilamide is a polymer made of acrylamide (C3H5NO) and bis-acrilamide (N,N’-methylene-bis-acrylamide C7H10N2O2) Acrylamide O CH CH2 NH2 C bis-Acrylamide O CH CH2 NH2 C Acrylamide Acrylamide

Polyacrylamide Gels Acrylamide polymerizes in the presence of free radicals typically supplied by ammonium persulfate O CH CH2 NH2 C O CH CH2 NH2 C SO4-.

Polyacrylamide Gels Acrylamide polymerizes in the presence of free radicals typically supplied by ammonium persulfate TMED (N,N,N’,N’-tetramethylethylenediamine) serves as a catalyst in the reaction NH2 O CH CH2 C O CH CH2 NH2 C SO4-.

Polyacrylamide Gels bis-Acrylamide polymerizes along with acrylamide forming cross-links between acrylamide chains O CH CH2 NH2 C O CH CH2 NH2 C bis-Acrylamide O CH CH2 NH2 C O CH CH2 NH2 C

Polyacrylamide Gels bis-Acrylamide polymerizes along with acrylamide forming cross-links between acrylamide chains

Polyacrylamide Gels Pore size in gels can be varied by varying the ratio of acrylamide to bis-acrylamide Protein separations typically use a 29:1 or 37.5:1 acrylamide to bis ratio Lots of bis-acrylamide Little bis-acrylamide

SDS-PAGE 3 2 1 Addition of SDS 1 2 3 2 3 1 Protein becomes rod-shaped with uniform charge distribution Addition of SDS

The End