Protein Full Length Sequencing

Full-Length Protein Sequencing refers to the determination of the complete amino acid sequence of a protein from the N-terminus to the C-terminus. This process involves a series of techniques, which are critical for understanding the biological function of proteins, studying the interactions between proteins, and developing new drugs. Common methods for full-length protein sequencing include Mass Spectrometry (MS), protein sequencers, and bioinformatics methods. Taking mass spectrometry as an example, it usually involves the following steps:

 

Analysis Workflow

1. Sample Preparation

Proteins are extracted from biological samples. This can be achieved through cell culture, tissue isolation, or other methods. The extracted proteins are then purified and separated through electrophoresis, liquid chromatography, or other separation methods to obtain a single protein sample.

 

2. Proteolytic Digestion

The separated protein sample is digested into small peptides using an enzyme (usually trypsin).

 

3. Mass Spectrometry Analysis

Mass spectrometry techniques, such as Tandem Mass Spectrometry (MS/MS), are used to analyze these peptides to determine their mass and amino acid sequence. This usually involves the use of a mass spectrometer and database searching to match the experimental data with known protein sequences.

 

4. Data Analysis and Assembly

The peptide sequence data obtained are analyzed and assembled to determine the full-length sequence of the protein.

 

5. Verification and Confirmation

Further experimental validation, such as Western blotting, re-analysis of mass spectrometry data, etc., is usually required to ensure the accuracy of the results.

 

Applications

Full-length protein sequencing has a wide range of applications in biomedical research, protein engineering, drug development, and disease diagnosis. Through full-length sequencing, researchers can better understand the function and structure of proteins, thus promoting the development of related fields.

 

1. Protein Function Studies

 Full-length sequencing helps us understand the exact function of a specific protein, including its catalytic activity, binding sites, active sites, and interactions with other proteins.

 

2. Biopharmaceuticals

In drug development, full-length protein sequencing is used to identify and verify the structure of recombinant proteins or monoclonal antibodies, ensuring their correctness and biological activity.

 

3. Protein Engineering and Biotechnology

With the information of the full-length protein sequence, proteins with specific properties or improved functions can be designed and constructed for industrial, research, or therapeutic applications.

 

4. Molecular Biology Research

By comparing protein sequences from different species or under different conditions, the evolutionary relationships, functional conservation, and adaptive changes of proteins can be revealed.

 

5. Disease Diagnosis and Biomarker Development

Full-length protein sequencing helps identify disease-related protein variants, providing information for disease diagnosis and the discovery of biomarkers.

 

6. Structural Biology

The full-length protein sequence is critical for the prediction and resolution of protein three-dimensional structure, helping to understand how proteins work at the molecular level.