Interpretation of Peptide Sequencing and Structural Modifications

An unknown peptide was isolated from the leaf of a medicinal plant and purified to homogeneity. The peptide did not yield any sequence when subjected to Edman degradation. However, tryptic digest of the peptide produced a unique sequence. The mass of the intact peptide was found to be 18 Da less than that obtained from the trypsin treated sample. The possible interpretation of the above experimental results could be that
P. The N-terminus of the peptide was blocked by acetylation or methylation.
Q. The peptide was cyclic and contained at least one internal arginine or lysine residue.
R. The peptide was cyclic and contained a lysine or arginine residue at the C-terminus.
S. The peptide was cyclized by peptide bond formation between a-amino group and a-carboxyl group. Which of the above statements is true?
A. p and S
C. Q and R
B. P and Q
D. Q and S

Understanding the structure and sequence of peptides is crucial in molecular biology and biochemistry. In peptide sequencing experiments, certain modifications or structural features can prevent accurate analysis. This article explains why an unknown peptide may resist Edman degradation and how tryptic digestion and mass spectrometry provide valuable insights into its structure.

Correct Answer: (B) P and Q

The most likely interpretation is that:

• The N-terminus of the peptide is blocked by acetylation or methylation (P).
• The peptide is cyclic and contains at least one internal arginine or lysine residue (Q).

Keyphrase: Peptide Sequencing Challenges

Peptide sequencing challenges often arise due to post-translational modifications and cyclic structures.

Understanding Peptide Sequencing

What is Peptide Sequencing?

Peptide sequencing is the process of determining the amino acid sequence of a peptide. Techniques like Edman degradation and mass spectrometry are commonly used for this purpose.

• Edman Degradation:

  • Sequential cleavage of amino acids from the N-terminal end of a peptide.
  • Only works when the N-terminal is free (unblocked).

• Mass Spectrometry:

  • Measures the mass-to-charge ratio of peptide fragments.
  • Helps identify modifications and cyclic structures.

Challenges in Peptide Sequencing

1. Blocked N-Terminus

• If the N-terminal of a peptide is blocked by acetylation or methylation, Edman degradation cannot initiate cleavage.
• This explains why the peptide did not yield a sequence using Edman degradation.

2. Cyclic Peptide Structure

• A cyclic peptide lacks a free N-terminal and C-terminal, which can interfere with Edman degradation.
• Trypsin can cleave at internal lysine or arginine residues, revealing the sequence of fragments.
• The increase in mass after tryptic digestion suggests the cleavage of a cyclic bond.

3. Mass Difference of 18 Da

• The mass difference of 18 Da corresponds to the loss of a water molecule (H2O), indicating that the peptide was originally cyclized.
• Opening of the cyclic bond during tryptic digestion would result in an increase in mass.

Possible Interpretations

Why P and Q are Correct

• The blocked N-terminal prevents Edman degradation.
• Trypsin’s ability to cleave the peptide indicates an internal lysine or arginine, supporting a cyclic structure.

Importance of Peptide Sequencing in Research

Peptide sequencing is critical for:

• Identifying protein modifications.
• Studying post-translational modifications.
• Developing peptide-based drugs.
• Understanding peptide-protein interactions.

Applications in Molecular Biology and Biotechnology

1. Drug Discovery

• Modified peptides are used as therapeutic agents.
• Cyclic peptides often have enhanced stability and bioavailability.

2. Structural Biology

• Cyclic peptides help model protein folding and interaction sites.

3. Protein Engineering

• Understanding peptide sequences allows for the design of modified proteins with specific functions.

Why Peptide Sequencing is Important for CSIR NET and Other Exams

Peptide sequencing and mass spectrometry are key topics in:

• CSIR NET Life Science
• IIT JAM
• GATE Biotechnology
• DBT JRF

Understanding these concepts will help you solve complex exam questions with accuracy.

Comparison of Peptide Sequencing Techniques

Challenges in Studying Cyclic Peptides

Conclusion

The inability of an unknown peptide to yield a sequence using Edman degradation points to an N-terminal modification and a cyclic structure. The mass increase of 18 Da after tryptic digestion supports this hypothesis. Peptide sequencing techniques, including Edman degradation, mass spectrometry, and trypsin digestion, are crucial for understanding protein structure and function.

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FAQs

Q1. Why did the peptide not yield a sequence with Edman degradation?
The N-terminal may have been blocked by acetylation or methylation.

Q2. What does an increase of 18 Da after tryptic digestion suggest?
It indicates the breaking of a cyclic bond and the addition of a water molecule.

Q3. How does trypsin help in peptide sequencing?
Trypsin cleaves at lysine and arginine residues, generating identifiable fragments.

This article was prepared with insights from Let’s Talk Academy, the top institute for life science competitive exams.