N-Terminal Sequencing Mass Spectrometry
N-terminal sequencing mass spectrometry is a technique used to determine the N-terminal amino acid sequence of proteins or peptides by employing mass spectrometric methods. In this technique, protein or peptide samples are ionized and then separated and detected in a mass spectrometer based on the mass-to-charge ratio (m/z) of the ions. Specific cleavage methods are typically utilized to systematically fragment the protein or peptide from the N-terminus into a series of fragment ions. By examining the mass-to-charge ratios of these fragment ions and the differences in their masses, the N-terminal amino acid sequence can be deduced. Common techniques such as electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI) convert proteins or peptides into gaseous ions, which are then separated and detected in a mass spectrometer.
N-terminal sequencing mass spectrometry allows us to identify the initial sequence of a protein segment, which is crucial for understanding protein function, investigating protein modifications, developing new pharmaceuticals, and diagnosing diseases. For example, in biopharmaceuticals, N-terminal sequencing verifies the accuracy of recombinant proteins, ensuring they meet drug production standards. In fundamental biological research, it aids in elucidating protein modification mechanisms, like acetylation and methylation, that occur at the N-terminus and influence protein stability and function. Additionally, N-terminal sequencing mass spectrometry is applied in clinical diagnostics to identify specific protein markers for diagnosing cancers or other diseases.
Main Steps of N-terminal Sequencing Mass Spectrometry Technology
1. Sample Preparation
Initially, target proteins or peptides are extracted and purified from biological samples to ensure sample purity and concentration are suitable for mass spectrometry analysis. This may involve techniques such as cell lysis, centrifugation, and chromatographic separation.
2. Enzymatic or Chemical Cleavage
To facilitate mass spectrometry analysis, proteins or peptides are typically cleaved into appropriately sized fragments. Common methods include enzymatic digestion with proteases like trypsin or chemical cleavage using reagents such as cyanogen bromide.
3. Mass Spectrometry Analysis
The processed sample is introduced into a mass spectrometer for analysis, generating a mass spectrum where each peak represents an ion with a specific mass-to-charge ratio.
4. Data Processing and Analysis
Specialized mass spectrometry data analysis software is used to identify, match, and interpret peaks in the mass spectrum. Based on the mass differences of the fragment ions, the N-terminal amino acid sequence is inferred. Comparison with protein databases can also be conducted to identify unknown proteins.
Advantages of N-terminal Sequencing Mass Spectrometry Technology
1. High Sensitivity
Capable of detecting protein or peptide samples at picomole to femtomole levels, significant for analyzing trace samples.
2. High Resolution
Allows precise measurement of ion mass-to-charge ratios, distinguishing ions with very small mass differences, thus accurately determining amino acid sequences.
3. Rapid and Efficient
A single analysis can quickly yield extensive information, enabling the rapid analysis of multiple samples.
4. No Labeling Required
Unlike some traditional sequencing methods, mass spectrometry analysis does not require radioactive or fluorescent labeling, simplifying the process.
MtoZ Biolabs offers professional N-terminal sequencing mass spectrometry services, dedicated to providing high-quality analytical results. Whether in drug development or biomarker research, our N-terminal sequencing mass spectrometry services can support your efforts. Through a customized service process, we ensure that each project meets specific client needs, providing strong support for your research and development.
MtoZ Biolabs, an integrated chromatography and mass spectrometry (MS) services provider.
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