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N-terminal Amino Acid Sequence Analysis

BOC Sciences provides N-terminal sequencing services based on mass spectrometry and non-mass spectrometry technologies, which can achieve accurate interpretation of N-terminal amino acid sequences and ensure coverage and accuracy of sequencing results.

Why N-terminal Sequencing?

The N-terminus is usually the starting point of protein or peptide synthesis. The composition of the N-terminal sequence has a huge impact on biological functions, such as protein half-life, protein localization in subcellular organelles, etc. N-terminal sequencing is to accurately analyze the amino acid species and sequence of the N-terminal. N-terminal sequencing helps to analyze the structure of protein or peptide molecules and reveal their biological functions. With the development of the modern pharmaceutical industry, a large number of protein and peptide drugs have emerged. The analysis and confirmation of the N-terminal sequences of these drug molecules is also an important part of quality control.

Our N-Terminal Sequencing Services

  • N-terminal Sequencing Based on Edman Degradation

We use the Edman degradation method to perform amino acid N-terminal sequencing services on samples provided by customers within 1-2 weeks, and can achieve the determination of 20-40 amino acids at a time. The sample state provided by the customer can be dry powder or solution. If the number of amino acids to be sequenced is relatively large, the total amount of the sample should be 2mg-3mg, the purity should be ≥95%, and the concentration should be ≥1mg/mL. It is necessary to ensure that the content of volatile inorganic salts < 20 mM and the content of non-volatile inorganic salts < 5 mM in the sample. After sequencing is complete, we deliver a full project report.

    • Edman Degradation Sequencing Analysis Steps

(1) Under alkaline conditions, Phenylisothiocyanate (PITC) binds to the N-terminal amino group of the protein or peptide to be tested.

(2) Selectively excise N-terminal residues in acidic solution.

(3) The residues bound by PITC are transformed into more stable phenylthiohydantoin (PTH) residues. The generated PTH-amino acids were analyzed by HPLC, and the amino acid species were identified according to the elution time.

Repeating the amino acid identification reaction can achieve the analysis of the amino acid sequence of the N-terminal amino acid in the direction from the N-terminal to the C-terminal.

    • Advantages of Edman Degradation Sequencing Service

(1) It can accurately analyze the N-terminal amino acid sequence, especially can distinguish isoleucine, leucine, glutamine and lysine.

(2) Amino acid information can be directly analyzed to identify unknown proteins.

(3) Reproducible determination of nascent N-terminal amino acids.

(4) Fast response time.

N-terminal Sequencing Based on Edman Degradation

  • N-terminal Sequencing Based on Mass Spectrometry

The N-terminal sequencing technology based on mass spectrometry can realize the determination of the amino acid sequence blocked at the N-terminus, which is complementary to Edman degradation sequencing. We use mass spectrometry technology to perform N-terminal sequencing services on samples provided by customers within a certain period. After sequencing is complete, we deliver mass spec files and coverage maps.

    • Advantages of Mass Spectrometry Sequencing Service

(1) The amino acid sequence of unknown peptides can be determined.

(2) Non-standard amino acids can be identified.

(3) It is not restricted by the N-terminal block, and can position the post-translational processing or chemical modification of the side chain group.

(4) The sensitivity has been improved.

    • Mass Spectrometry Sequencing Service Process

(1) Use an enzyme to digest the target protein.

(2) N-terminal sequence analysis by mass spectrometry.

(3) Compare the spectrum obtained by mass spectrometry with the theoretical sequence.

(4) Complete the result report.

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