Top-down proteomics.
David S RobertsJoseph A LooYury O TsybinXiaowen LiuSi WuJulia Chamot-RookeJeffrey N AgarLjiljana Pasa-TolicLloyd M SmithYing GePublished in: Nature reviews. Methods primers (2024)
Proteoforms, which arise from post-translational modifications, genetic polymorphisms and RNA splice variants, play a pivotal role as drivers in biology. Understanding proteoforms is essential to unravel the intricacies of biological systems and bridge the gap between genotypes and phenotypes. By analysing whole proteins without digestion, top-down proteomics (TDP) provides a holistic view of the proteome and can decipher protein function, uncover disease mechanisms and advance precision medicine. This Primer explores TDP, including the underlying principles, recent advances and an outlook on the future. The experimental section discusses instrumentation, sample preparation, intact protein separation, tandem mass spectrometry techniques and data collection. The results section looks at how to decipher raw data, visualize intact protein spectra and unravel data analysis. Additionally, proteoform identification, characterization and quantification are summarized, alongside approaches for statistical analysis. Various applications are described, including the human proteoform project and biomedical, biopharmaceutical and clinical sciences. These are complemented by discussions on measurement reproducibility, limitations and a forward-looking perspective that outlines areas where the field can advance, including potential future applications.
Keyphrases
- data analysis
- tandem mass spectrometry
- liquid chromatography
- mass spectrometry
- ultra high performance liquid chromatography
- high performance liquid chromatography
- endothelial cells
- protein protein
- amino acid
- electronic health record
- current status
- gas chromatography
- simultaneous determination
- big data
- high resolution
- binding protein
- small molecule
- amyotrophic lateral sclerosis
- dna methylation
- risk assessment
- genome wide
- deep learning
- artificial intelligence
- induced pluripotent stem cells
- human health
- nucleic acid