The emerging landscape of single-molecule protein sequencing technologies

Alfaro, Javier A.; Bohländer, Peggy R.; Dai, Mingjie; Filius, Mike; Howard, Cecil J.; Kooten, Xander F. van; Ohayon, Shilo; Pomorski, Adam; Schmid, Sonja; Aksimentiev, Aleksei; Anslyn, Eric V.; Bedran, Georges; Cao, Chunhua; Chinappi, Mauro; Coyaud, Étienne; Dekker, Cees; Dittmar, Gunnar; Drachman, Nicholas; Eelkema, Rienk; Goodlett, David R.; Hentz, Sébastien; Kalathiya, Umesh; Kelleher, Neil L.; Kelly, Ryan; Kelman, Zvi; Kim, Sung Hyun; Küster, Bernhard; Rodríguez-Larrea, David; Lindsay, Stuart; Maglia, Giovanni; Marcotte, Edward M.; Marino, John P.; Masselon, Christophe; Mayer, Michael; Samaras, Patroklos; Sarthak, Kumar; Sepiashvili, Lusia; Stein, Derek; Wanunu, Meni; Wilhelm, Mathias; Yin, Peng; Meller, A.; Joo, Chirlmin

doi:10.1038/s41592-021-01143-1

Cited by 245 publications

(246 citation statements)

References 140 publications

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“…The resulting unbiased and holistic view of both molecular (e.g., gene expression) and cellular (i.e., single cell) dimensions enables identification of new cell (sub) populations, and those that associate with pathogenesis will be highlighted and further explored. With rapid development of single cell proteomics [ 229 – 231 ], single cell type proteomics [ 232 , 233 ], and single-molecule protein sequencing technologies [ 234 ], the application to the AD field is expected in the near future.…”

Section: Conclusion and Perspectives Of Ad Proteomics-based Systems Biologymentioning

confidence: 99%

Proteomic landscape of Alzheimer’s Disease: novel insights into pathogenesis and biomarker discovery

Bai

Vanderwall

et al. 2021

Mol Neurodegeneration

130

100

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Mass spectrometry-based proteomics empowers deep profiling of proteome and protein posttranslational modifications (PTMs) in Alzheimer’s disease (AD). Here we review the advances and limitations in historic and recent AD proteomic research. Complementary to genetic mapping, proteomic studies not only validate canonical amyloid and tau pathways, but also uncover novel components in broad protein networks, such as RNA splicing, development, immunity, membrane transport, lipid metabolism, synaptic function, and mitochondrial activity. Meta-analysis of seven deep datasets reveals 2,698 differentially expressed (DE) proteins in the landscape of AD brain proteome (n = 12,017 proteins/genes), covering 35 reported AD genes and risk loci. The DE proteins contain cellular markers enriched in neurons, microglia, astrocytes, oligodendrocytes, and epithelial cells, supporting the involvement of diverse cell types in AD pathology. We discuss the hypothesized protective or detrimental roles of selected DE proteins, emphasizing top proteins in “amyloidome” (all biomolecules in amyloid plaques) and disease progression. Comprehensive PTM analysis represents another layer of molecular events in AD. In particular, tau PTMs are correlated with disease stages and indicate the heterogeneity of individual AD patients. Moreover, the unprecedented proteomic coverage of biofluids, such as cerebrospinal fluid and serum, procures novel putative AD biomarkers through meta-analysis. Thus, proteomics-driven systems biology presents a new frontier to link genotype, proteotype, and phenotype, accelerating the development of improved AD models and treatment strategies.

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Section: Conclusion and Perspectives Of Ad Proteomics-based Systems Biologymentioning

confidence: 99%

Proteomic landscape of Alzheimer’s Disease: novel insights into pathogenesis and biomarker discovery

Bai

Vanderwall

et al. 2021

Mol Neurodegeneration

130

100

View full text Add to dashboard Cite

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“…Until this is done, we will be unable to advance our methods or thus deep and routine proteomic analyses to the extent that is both necessary and sufficient for unbiased identification of genuine highly selective biomarkers and drug targets. That said, there are indeed critical ongoing efforts to improve both sample and data analyses, and the best of these must be integrated into a continuously developing unified approach to proteoform and thus proteome analysis [29,81,88,[257][258][259][260][261][262][263][264][265][266][267][268][269][270][271][272][273][274]. We are hopeful and confident of a more collaborative and unbiased future for the discipline of proteomics.…”

Section: What Next?mentioning

confidence: 99%

Proteomes Are of Proteoforms: Embracing the Complexity

2021

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Proteomes are complex—much more so than genomes or transcriptomes. Thus, simplifying their analysis does not simplify the issue. Proteomes are of proteoforms, not canonical proteins. While having a catalogue of amino acid sequences provides invaluable information, this is the Proteome-lite. To dissect biological mechanisms and identify critical biomarkers/drug targets, we must assess the myriad of proteoforms that arise at any point before, after, and between translation and transcription (e.g., isoforms, splice variants, and post-translational modifications [PTM]), as well as newly defined species. There are numerous analytical methods currently used to address proteome depth and here we critically evaluate these in terms of the current ‘state-of-the-field’. We thus discuss both pros and cons of available approaches and where improvements or refinements are needed to quantitatively characterize proteomes. To enable a next-generation approach, we suggest that advances lie in transdisciplinarity via integration of current proteomic methods to yield a unified discipline that capitalizes on the strongest qualities of each. Such a necessary (if not revolutionary) shift cannot be accomplished by a continued primary focus on proteo-genomics/-transcriptomics. We must embrace the complexity. Yes, these are the hard questions, and this will not be easy…but where is the fun in easy?

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“…Nanopore sequencing is an interdisciplinary technology where a wide variety of fields intersect, from protein science and biophysics to chemistry and computer science ( Alfaro et al., 2021 ; Robertson et al., 2021 ). As the translocation methods discussed above involve the conjugation of translocation-assisting molecules (e.g., oligonucleotides and peptides) to a protein terminus, protein modification and conjugation chemistry are a key step to allow analysis of native proteins, analogous to how nanopore DNA/RNA sequencing requires sample preparation steps such as adapter ligation ( Garalde et al., 2018 ; Jain et al., 2016 ).…”

Section: Site-specific Protein Modification and Conjugation Chemistrymentioning

confidence: 99%

“…To address these challenges, complementary or potentially disruptive platforms for next-generation protein analysis and sequencing have been envisioned recently ( Alfaro et al., 2021 ; Asandei et al., 2020 ; Cressiot et al., 2020 ; Hu et al., 2021 ; Restrepo-Pérez et al., 2018 ; Timp and Timp, 2020 ). These emerging techniques include tunneling currents, single-molecule fluorescence, and nanopores.…”

Section: Introductionmentioning

confidence: 99%

Herding cats: Label-based approaches in protein translocation through nanopore sensors for single-molecule protein sequence analysis

2021

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Summary Proteins carry out life's essential functions. Comprehensive proteome analysis technologies are thus required for a full understanding of the operating principles of biological systems. While current proteomics techniques suffer from limitations in sensitivity and/or throughput, nanopore technology has the potential to enable de novo protein identification through single-molecule sequencing. However, a significant barrier to achieving this goal is controlling protein/peptide translocation through the nanopore sensor for processive strand analysis. Here, we review recent approaches that use a range of techniques, from oligonucleotide conjugation to molecular motors, aimed at driving protein strands and peptides through protein nanopores. We further discuss site-specific protein conjugation chemistry that could be combined with these translocation approaches as future directions to achieve single-molecule protein detection and sequencing of native proteins.

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The emerging landscape of single-molecule protein sequencing technologies

Cited by 245 publications

References 140 publications

Proteomic landscape of Alzheimer’s Disease: novel insights into pathogenesis and biomarker discovery

Proteomic landscape of Alzheimer’s Disease: novel insights into pathogenesis and biomarker discovery

Proteomes Are of Proteoforms: Embracing the Complexity

Herding cats: Label-based approaches in protein translocation through nanopore sensors for single-molecule protein sequence analysis

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