Principle of Protein Structure Identification

Proteins, as fundamental functional units of living organisms, are essential in understanding biological processes and developing new drugs. Protein structure determination is a key approach to elucidate protein functions. This article will detail the principles and methods of protein structure determination.

 

Levels of Protein Structure

Protein structure is typically divided into four levels: primary, secondary, tertiary, and quaternary structures.

 

1. Primary Structure

This refers to the amino acid sequence of the protein, the most basic level of protein structure. Determining the amino acid sequence provides a foundation for further structural analysis.

 

2. Secondary Structure

This refers to local, regular folding patterns within the protein chain, primarily including α-helices and β-sheets, which are stabilized by hydrogen bonds.

 

3. Tertiary Structure

This describes the overall three-dimensional folding of the entire protein molecule, reflecting its overall shape.

 

4. Quaternary Structure

This refers to the assembly of multiple protein subunits into a larger complex, held together by non-covalent bonds.

 

Main Methods of Protein Structure Determination

There are several methods for protein structure determination, with X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy, and cryo-electron microscopy (Cryo-EM) being the most common.

 

1. X-Ray Crystallography

(1) Principle: This technique relies on the diffraction of X-rays by protein crystals. The resulting diffraction pattern is used to derive an electron density map, which, combined with the amino acid sequence, allows for the construction of a three-dimensional model of the protein.

(2) Advantages: High resolution; suitable for most stable, easily crystallized proteins.

(3) Limitations: Requires high-quality protein crystals, which are difficult to obtain for some proteins.

 

2. Nuclear Magnetic Resonance (NMR) Spectroscopy

(1) Principle: This method is based on the absorption and emission of electromagnetic radiation by atomic nuclei in a magnetic field. By analyzing NMR signals, the three-dimensional structure of the protein in solution can be inferred.

(2) Advantages: Can study protein dynamics under near-physiological conditions; suitable for small proteins and protein complexes.

(3) Limitations: Signal overlap becomes severe for large molecules or complex systems, making interpretation difficult.

 

3. Cryo-Electron Microscopy (Cryo-EM)

(1) Principle: Protein samples are rapidly frozen to liquid nitrogen temperatures, and imaged using an electron microscope at low temperatures. Three-dimensional reconstruction from numerous single-particle images provides structural information.

(2) Advantages: Does not require crystallization; suitable for large molecular complexes and membrane proteins.

(3) Limitations: Expensive equipment, complex data processing, and relatively lower resolution.

 

In summary, protein structure determination is a vital tool in biological research. Techniques such as X-ray crystallography, NMR spectroscopy, and cryo-EM allow scientists to resolve protein structures, thus revealing their functions and providing crucial information for biomedical research and applications.