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SDS-PAGE Sodium Dodecyl Sulfate - Polyacrylamid Gel Electrophoresis. Sodium Dodecyl Sulfate. SDS is a common ingredient in detergents Other names for SDS include laurel sulfate and sodium laurel sulfate As a detergent SDS destroys protein secondary, tertiary and quaternary structure

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sodium dodecyl sulfate
Sodium Dodecyl Sulfate
  • SDS is a common ingredient in detergents
  • Other names for SDS include laurel sulfate and sodium laurel 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

O

H-C-C-C-C-C-C-C-C-C-C-C-C-O-S-O-Na+

O

Non-polar

Hydrophobic tail

Polar

Hydrophilic head

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

H

SDSSodium Dodecyl Sulfate

C12H25NaO4S

  • Because it is amphipathic, SDS is a potent detergent
sds and proteins1
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

O

Acrylamide

C

NH2

CH2

CH

CH2

CH

O

C

NH2

CH2

Acrylamide

Acrylamide

O

C

NH2

bis-Acrylamide

CH2

CH

Polyacrylamide Gels
  • Polyacrilamide is a polymer made of acrylamide (C3H5NO) and bis-acrilamide (N,N’-methylene-bis-acrylamide C7H10N2O2)
polyacrylamide gels1

O

O

C

C

NH2

NH2

CH2

CH2

CH

CH

SO4-.

Polyacrylamide Gels
  • Acrylamide polymerizes in the presence of free radicals typically supplied by ammonium persulfate
polyacrylamide gels2

O

O

O

O

C

NH2

C

NH2

C

NH2

C

NH2

CH2

CH

CH2

CH

CH2

CH

CH2

CH

SO4-.

Polyacrylamide Gels

1. Acrylamide polymerizes in the presence of free radicals typically supplied by ammonium persulfate

  • 2.TMED (N,N,N’,N’-tetramethylethylenediamine) serves as a catalyst in the reaction
polyacrylamide gels3

O

O

O

C

NH2

C

NH2

C

NH2

CH2

CH

CH2

CH

CH2

CH

CH2

CH

O

O

O

C

NH2

C

NH2

C

NH2

CH2

O

CH2

CH

CH2

CH

O

C

NH2

C

NH2

CH2

CH

CH2

CH

bis-Acrylamide

Polyacrylamide Gels
  • bis-Acrylamide polymerizes along with acrylamide forming cross-links between acrylamide chains
polyacrylamide gels4
Polyacrylamide Gels
  • bis-Acrylamide polymerizes along with acrylamide forming cross-links between acrylamide chains
polyacrylamide gels5

Lots of bis-acrylamide

Little bis-acrylamide

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

3

Addition of SDS

2

Protein becomes rod-shaped with uniform charge distribution

1

SDS-PAGE

1 2 3

slide13

The

End