Mass Spectrometry-Based Protein Modification Sites Analysis Service

Post-translational modifications (PTMs) of proteins play a vital regulatory role in biological processes. By adding chemical groups to specific residues, PTMs can significantly affect a protein’s conformation, stability, subcellular localization, and interaction capabilities. These modifications are key to regulating signal transduction, the cell cycle, stress responses, and metabolic reprogramming.

 

Common PTMs include phosphorylation, acetylation, methylation, ubiquitination, and glycosylation. In recent years, emerging modifications such as lactylation, hydroxybutyrylation, and N-acetylglutamylation have further expanded the functional landscape of PTMs. As these modifications typically occur at low abundance and specific sites, traditional analytical methods are often inadequate for accurate detection and localization. High-sensitivity, high-throughput detection technologies have thus become essential tools for PTM research.

 



Zhong, Q. et al. MedComm (2020). 2023.

Figure 1. Common Types of Protein Posttranslational Modifications

 

Leveraging advanced high-resolution mass spectrometry platforms, MtoZ Biolabs offers a comprehensive Mass Spectrometry-Based Protein Modification Sites Analysis Service designed to support the accurate identification of various PTM sites, enabling researchers to investigate protein function regulation and discover potential disease-related biomarkers.

 

Services at MtoZ Biolabs

1. Target Protein Modification Site Analysis

MtoZ Biolabs provides Targeted Protein Modification Site Analysis focused on the precise detection and characterization of modification sites on specific proteins. Using enzymatic digestion, antibody- or chemistry-based enrichment, and high-resolution LC-MS/MS analysis, we accurately localize modification sites, determine modification-specific mass shifts, and evaluate site-level microheterogeneity.

This service is ideal for validating modification events on recombinant or native proteins and supports structural characterization, functional studies, and mechanistic evaluation in basic and translational research.

2. Proteome-Wide Modification Site Analysis

MtoZ Biolabs' Proteome-Wide Modification Site Analysis enables large-scale, site-specific profiling of post-translational modifications across complex biological samples. By integrating enrichment strategies such as antibody affinity, IMAC, TiO₂, chemical labeling, or resin capture with multidimensional LC-MS/MS and advanced fragmentation modes including HCD, ETD, and EThcD, we systematically identify and quantify thousands of modification sites.

Supported by comprehensive bioinformatics analysis, this service reveals PTM distribution, dynamic regulation, pathway associations, and biological relevance, providing deep insights into signaling networks, protein regulation, and disease-related modification mechanisms.

Analysis Workflow



Dephoure, N. et al. Mol Biol Cell. 2013.

Figure 2. Tandem Mass Spectrometry (MS/MS) Analysis of Protein Phosphorylation Site

Service Advantages

1. Stable and Reliable Platform: High-end mass spectrometers and standardized workflows ensure reproducibility and high accuracy.

2. Comprehensive PTM Coverage: Detection of both classical and novel protein modifications, including phosphorylation, acetylation, ubiquitination, methylation, glycosylation, lipidation, SUMOylation, NEDDylation, UFMylation, lactylation, succinylation, and other emerging regulatory modifications.

3. Flexible Sample Compatibility: Suitable for various sample types, including cells, tissues, fluids, and recombinant proteins.

4. Customizable Analysis Depth: Tailored for qualitative or quantitative analysis with optional visualized bioinformatics reports.

5. Full Project Support: End-to-end technical assistance, from experimental design to data interpretation, for diverse research needs.

 

Applications

Protein modification site analysis is widely applicable in fundamental research, disease mechanism studies, and drug development, including:

1. Signal Transduction Studies: Monitor PTM changes in key proteins under activation or inhibition.

2. Cancer Mechanism Research: Explore PTM-related pathways in tumor progression.

3. Target Validation and Screening: Evaluate drug impact on PTM status of candidate proteins.

4. Biomarker Discovery: Identify site-specific PTMs associated with disease progression.

5. Functional Studies: Verify how specific PTMs affect protein structure, activity, or interactions using mutation-based experiments.

 

Case Study

1. Characterization of Site-Specific N- and O-Glycopeptides from Recombinant Spike and ACE2 Glycoproteins Using LC-MS/MS Analysis

This study employed high-resolution LC-MS/MS to systematically characterize the site-specific N- and O-glycosylation profiles of recombinant SARS-CoV-2 spike protein (RBD and S2 subunit) and ACE2 proteins. A total of 148 N- and 28 O-glycopeptides were identified from RBD, 71 N-glycopeptides from the S2 subunit, and 139 N-glycopeptides from ACE2. Notably, novel glycan modifications including mannose-6-phosphate, GlcNAc-1-phosphate-6-O-mannose on N-glycans, and O-acetylation on O-glycans of RBD were reported for the first time. The study also revealed dominant glycosylation sites and glycan types depending on protein origin and expression system. Mass Spectrometry-Based Protein Modification Sites Analysis service enables high-coverage identification of diverse glycosylation types and PTMs at the site-specific level, supporting detailed structural profiling of recombinant or native proteins in viral, therapeutic, or host interaction studies.

 



Song, JH. et al. Int J Mol Sci. 2024.

Figure 3. Site-Specific Characterization of O-Glycosylation and O-Acetylation on the RBD-Derived Glycopeptide VQQPTESIVR by EThcD-MS/MS

 

2. Mass Spectrometric Identification of Novel Lysine Acetylation Sites in Huntingtin

This study employed high-resolution mass spectrometry to systematically identify lysine acetylation sites in the N-terminal fragment of the Huntingtin protein (Htt23Q 1–612). Through immunoprecipitation, SDS-PAGE separation, enzymatic digestion, and LC-MS/MS analysis, five acetylation sites were identified, including three novel ones: Lys-178, Lys-236, and Lys-345. Additionally, a multiple reaction monitoring (MRM) assay was developed to compare the acetylation level of Lys-178 between wild-type Htt23Q and mutant Htt148Q, and the presence of several acetylation sites was confirmed using site-specific antibodies. Mass Spectrometry-Based Protein Modification Sites Analysis service enables systematic identification and validation of lysine acetylation and other post-translational modifications, supporting quantitative comparisons between wild-type and mutant proteins in disease-related models.

 



Cong, X. et al. Mol Cell Proteomics. 2011. 

Figure 4. Three Novel Htt Acetylation Sites Identified by MS

 

FAQ

Q1: How much sample is required?

A1: We recommend a minimum of 100 µg of total protein. Specific enrichment strategies may have different requirements. Detailed sample preparation guidelines are available upon request.

 

Q2: Do you offer quantitative analysis?

A2: Yes. We support both labeled quantification methods (e.g., TMT, iTRAQ) and label-free approaches for relative quantification.

 

Q3: What is the typical turnaround time?

A3: Standard turnaround is approximately 3–4 weeks, depending on project complexity and sample number.

 

Q4: Are the data suitable for publication?

A4: Yes. We provide data files and reports that meet the standards of major scientific journals, supporting submission, funding applications, and academic defense.

 

If you are interested in exploring protein modification sites, contact MtoZ Biolabs to learn more. Our team is committed to providing high-quality, efficient Mass Spectrometry-Based Protein Modification Sites Analysis Service to support your scientific advancement.