Advances in protein sequencing: Techniques, challenges and prospects

Proteins are major building blocks of life, which are the principal undertakers of life activities. The protein components and distribution of a cell or an organ can give us lots of key information to understand the biological process in human life as well as diagnose diseases. Protein sequencing can determine individual proteins’ sequences, structures, and post-translational modifications, deliver detailed insights into protein-protein interactions and functions, which lead to advances such as characterization of disease biomarkers and personalized medicine, transforming our comprehension of complex biological systems. While conventional methods such as Edman degradation and mass spectrometry are being enhanced by integrating with other techniques, emerging sequencing approaches such as DNA point accumulation in nanoscale topology (DNA-PAINT), recognition tunneling and nanopore sequencing have advanced significantly, moving beyond the proof-of-concept stage and achieving notable progress recently. This review will examine these techniques, discuss the advancements and inherent challenges, such as sensitivity, sample complexity, and low abundance of protein sample, and propose some corresponding solutions. Finally, a forward-looking perspective on the future of protein sequencing, emphasizing the need for more sensitive and high-throughput methods, the integration of multi-omics data, the role of computational approaches, and the expansion of protein sequencing into new application domains will also be presented. This review will serve as a valuable resource for researchers and practitioners, guiding future developments and applications in protein sequencing.