Advancements in MS-Based Techniques
Recent advancements in MS-based protein interactomics have significantly enhanced our understanding of PPIs.

Various techniques have been developed, each with its own set of advantages and limitations. Key approaches include:

Affinity Purification Mass Spectrometry (AP-MS)
AP-MS is one of the most widely used methods for mapping protein interactions on a large scale. This technique involves fusing a protein of interest (POI) with an affinity tag and expressing it in cells. Following cell lysis, the POI and its interacting proteins are purified using an affinity resin. The versatility of AP-MS is expanded by the diverse range of affinity tags available, such as GFP, FLAG, and His-tags. Advancements in MS technology have further improved the accuracy of PPI detection, allowing for a deeper understanding of cellular functions.

Cross-Linking Mass Spectrometry (XL-MS)
XL-MS is a powerful tool for deciphering the structures of protein complexes by stabilizing interactions through cross-linkers. This technique can differentiate direct from indirect interactions and provide detailed structural information. Recent methodologies include in vivo crosslinking and performing crosslinking in lysates to enhance the detection of transient or weak PPIs. Innovations such as APEX-based proximity labeling combined with XL-MS have expanded its capabilities to study dynamic interactomes within cells.

Proximity Labeling (PL)
PL techniques, such as BioID and APEX, offer complementary approaches to AP-MS by capturing transient interactions in living cells. BioID uses a mutant biotin ligase to label proteins near the POI, which are then purified using streptavidin beads. APEX, derived from ascorbate peroxidase, labels proteins via biotin-phenol in the presence of hydrogen peroxide. These methods have been applied to study a variety of cellular structures and processes, providing valuable insights into the dynamic nature of PPIs.

Challenges and Future Prospects
Despite the significant advancements in MS-based PPI mapping, challenges remain. The low efficiency of cross-linking reagents and the complexity of peptide mixtures can hinder comprehensive analysis. Future improvements in enrichment strategies, cross-linker design, and bioinformatics tools are essential for overcoming these challenges. Additionally, integrating MS-based techniques with other cutting-edge approaches, such as CRISPR-mediated genome editing and real-time imaging, holds great promise for advancing our understanding of the interactome.

Conclusion
MS-based approaches are transforming our knowledge of PPIs, offering detailed insights into the intricate networks of protein interactions within cells. These advancements are critical for elucidating cellular functions and understanding disease mechanisms. As the field continues to evolve, MS-based protein interactomics will undoubtedly play a pivotal role in unraveling the complexities of the proteome and advancing biomedical research.

For a detailed review, see the full recent article in Mass Spectrometry Reviews.

https://analyticalsciencejournals.onlinelibrary.wiley.com/doi/full/10.1002/mas.21887