In addition to providing bottom-up and top-down mass spectrometry (MS)-based proteomics workflows for post-translational modification (PTM) analysis, Creative Proteomics also provides a middle-down approach for the investigation of PTMs. As a compromise between bottom-up and top-down proteomics, this method is commonly used for polypeptide analysis. And these polypeptides retain more information about the coexisting PTMs and facilitate the analysis of combinatorial PTMs.
Middle-down MS-based proteomics
MS-based proteomics PTM analysis includes PTM site identification, PTM peptide quantification, PTM site occupancy analysis, among others. Different proteomics analysis techniques and analysis strategies can be used depending on the content of the analysis. The middle-down approach is a compromise between bottom-up and top-down proteomics, working with 5-20 kDa large polypeptides. In this strategy, proteins are usually digested using AspN and GluC instead of trypsin to obtain large peptides. The peptides are then sequenced using tandem mass spectrometry (MS/MS) techniques. The middle-down MS-based approach has two main advantages, including preserving more context for the PTMs and can be used in conjunction with an online separation policy to increase the number of recognized modification forms.
Features of middle-down proteomics
- High-throughput and feasible for polypeptide analysis.
- Protein and peptide fractionation dependent on size
- Work with 5-20 kDa large polypeptides produced by restricted proteolytic digestion.
- Suitable for the quantification of combinatorial PTMs.
Middle-down MS-based post-translational modification analysis service
We are a leading contract research organization (CRO) company focusing on MS-based PTM analysis worldwide. To meet the need of our customers to further investigate more PTMs and combinatorial PTMs, we offer a middle-down MS-based PTM analysis service. Our service includes experimental consultation and design, sample preparation for MS analysis, protein fractionation, restricted proteolysis, peptide fractionation, MS analysis, and data evaluation. We have experienced middle-down proteomics scientists and technical staff to provide you with quality PTM analysis services. We focus on the restricted digestion steps and optimized conditions for protein hydrolysis and use enzyme-assisted digestion and chemical-mediated proteolysis.
Application of our service
- Histone PTM analysis and identification
- Monoclonal antibody characterization
- Other therapeutic protein analysis
Major benefits of our service
- Advanced proteomics platform to ensure deep coverage.
- Experienced professionals work with the highest standards.
- Providing robust and reproducible results for future follow-up studies.
- Detailed analysis process and easy-to-follow reports.
Related services
Notably, to achieve comprehensive protein PTM analysis, the middle-down method can be combined with the top-down approach, namely, the MS/MS experiments are performed on middle-down sized proteolytic peptides and intact proteins. Creative Proteomics is dedicated to providing the optimal analysis strategy and solution according to the specific project requirements of our clients. Feel free to contact us for more information about our MS-based PTM analysis services. We are always available for your inquiries!
Reference
- Pandeswari, P. B., & Sabareesh, V. (2019). Middle-down approach: a choice to sequence and characterize proteins/proteomes by mass spectrometry. RSC advances, 9(1), 313-344.
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