Differences between SDS PAGE and Native PAGE

The difference between SDS-PAGE and native PAGE lies in how proteins are separated in the polyacrylamide gel. In SDS-PAGE, the migration rate of the protein is determined by its molecular weight only, whereas in native PAGE, the migration rate depends on both the size and overall charge of the protein. In this article, we will study the process of SDS PAGE and Native PAGE as well as the similarities and differences between SDS-PAGE and native PAGE.

Differences Between SDS PAGE and Native PAGE

The native page vs SDS page is discussed in the following table:

Criteria	SDS PAGE	Native PAGE
Description	A technique that separates the proteins based on their molecular weight or mass	A technique that separates the proteins based on their size, charge, and shape
Feature of Gel	Denaturing gel is used	Non-denaturing gel is used
Presence of SDS in Gel	SDS is present	SDS is absent
Buffer composition	The buffer has a reducing agent like DTT or BME	The buffer has no reducing agent
Sample preparation	Protein samples are heated	Protein samples are not heated
Separation criteria	Proteins are separated based on molecular weight	Proteins are separated based on molecular size and overall charge
Net charge on proteins	Always negative	Can either be positive or negative
Temperature	SDS PAGE is run at room temperature	Native PAGE is run at 4oC
Protein stability	Unstable	Stable
State of protein	Denatured	Native conformation or folded state
Protein recovery	Cannot be recovered post separation	Can be recovered post separation
Protein function	Proteins lose their function	Proteins retain their function
Used for	To detect presence of a protein; determine molecular weight (MW) of a protein; check protein expression etc.	To study the structure, subunit composition and function of proteins; purify proteins from a mixture in native form.
Ease of use	Easier, frequently used	Comparatively difficult, infrequent usage

What is SDS PAGE?

SDS-PAGE full form is Sodium dodecyl-sulfate polyacrylamide gel electrophoresis. It is a polyacrylamide gel electrophoresis technique, widely used in molecular biology. This technique is used to separate proteins by their molecular weight. It was developed by Ulrich K. Laemmli.

The SDS-PAGE electrophoresis uses sodium dodecyl sulphate (SDS), an anionic detergent, that binds to the hydrophobic regions of the proteins and denatures their three-dimensional structure. This is why the gel used in the SDS-PAGE method is called denaturing gel. Since proteins are denatured in gel, they cannot be retrieved in their functional form, post run. The buffers used in SDS PAGE have both denaturing and reducing agents. This method is widely used for determining molecular weight (MW) of a protein, purification and studying expression levels.

What is Native PAGE?

Native PAGE electrophoresis is another polyacrylamide gel electrophoresis technique, used in molecular biology to study the composition, native structure and function of the proteins. This technique separates the protein molecules based on the size and overall charge of the protein. It was developed by Ornstein and Davis. Since SDS is not used as a denaturing agent in the gel, the protein molecules retain their folded state and functionality. The buffers used in SDS PAGE do not contain any denaturing and reducing agents.

There are mainly two types of native PAGE:

• Blue Native PAGE (BN-PAGE): Here Coomassie brilliant blue dye is used during protein separation.
• Clear Native PAGE (CN-PAGE): Here proteins are separated based on their complex charge in a gradient gel without using any dye.

Similarities between SDS PAGE and Native PAGE

Some similarities between SDS PAGE and Native PAGE are:

• Gel Matrix: Both SDS-PAGE and Native PAGE use polyacrylamide gels for protein separation.
• Principle: Both techniques rely on electrophoresis, where proteins migrate through the gel matrix in response to an electric field.
• Electric Field: In both methods, an electric field is applied to drive the migration of charged proteins.
• Separation Basis: Both aim to separate proteins based on size, with smaller proteins moving faster through the gel.
• Buffer Usage: Both techniques use electrophoresis buffers to provide the necessary ionic environment for protein migration.
• Staining: After electrophoresis, proteins in both gels are visualized by staining for analysis.
• Molecular Biology Applications: Both are widely employed in molecular biology for protein analysis, purification, and molecular weight determination.
• Sample Loading: Proteins are loaded into wells at the top of the gel in both SDS-PAGE and Native PAGE.
• Migration Direction: Proteins migrate from the negative to the positive electrode during electrophoresis in both techniques.
• Equipment: The basic setup, including gel electrophoresis apparatus and power supply, is similar for both methods

Conclusion – Differences Between SDS PAGE and Native PAGE

In conclusion, SDS-PAGE and Native PAGE are two distinct techniques in molecular biology for protein separation. This article discusses the comparision between SDS PAGE and Native PAGE. SDS-PAGE separates proteins by molecular weight in denaturing conditions. Native PAGE separates proteins based on size and charge under non-denaturing conditions. It also preserves native conformation and functionality of the proteins. SDS-PAGE is used for protein detection and molecular weight determination. Native PAGE is used for studying protein structure, composition, and function.

FAQs on SDS PAGE and Native PAGE

Why is SDS not used in Native PAGE?

Native PAGE is employed to examine the composition and native structure of proteins. Therefore, SDS is not used due to its disruptive effect on the 3D structure of the proteins.

What are the Different Types of Native PAGE?

There are 3 types of native PAGE: Blue native PAGE (BN PAGE), Clear native PAGE (CN PAGE) and Preparative native continuous PAGE (PNC PAGE).

Why is APS and TEMED used in PAGE?

In the preparation of a polyacrylamide gel, Ammonium Persulfate (APS) and Tetramethylenediamine (TEMED) both are used for polymerization of acrylamide and bisacrylamide monomers.

What does Native Mean in Gel Electrophoresis?

In gel electrophoresis, “native” means that inside the polyacrylamide gel, the proteins are allowed to retain their three-dimensional shape or native conformation.

What do you Mean by Denaturing Gel?

A denaturing gel is a type of gel used in electrophoresis where the proteins lose their native, three-dimensional structure due to the presence of denaturing agents such SDS.

What is the Difference Between Native PAGE and Denaturing PAGE?

Native PAGE separates proteins based on their native structure, while denaturing PAGE separates proteins based on their molecular weight after denaturation.

What is the Difference Between Western and SDS-PAGE?

Western blotting is a technique used to detect specific proteins in a sample, while SDS-PAGE separates proteins based on their molecular weight.

What is the Difference Between SDS-PAGE and Electrophoresis?

SDS-PAGE is a type of electrophoresis technique specifically designed to separate proteins based on their molecular weight using SDS detergent, whereas electrophoresis is a broader term comprising of various techniques to separate molecules based on their charge and size using an electric field.

What is the Purpose of SDS in SDS-PAGE?

SDS in SDS-PAGE serves to denature proteins, bind to them, and confer a negative charge proportional to the protein’s mass, enabling separation based on size during electrophoresis.

Why is Beta Mercaptoethanol used in SDS-PAGE?

Beta-mercaptoethanol is used in SDS-PAGE to reduce disulfide bonds in proteins, thereby denaturing them and allowing for more accurate separation based on molecular weight during electrophoresis.

Why is Glycerol used in SDS-PAGE?

Glycerol in SDS-PAGE increases sample density, helping loading into gel wells for accurate protein separation by molecular weight.