Ribosomal Peptides and Small Proteins on the Rise

Scheidler, Christopher M.; Kick, Leonhard M.; Schneider, Sabine

doi:10.1002/cbic.201800715

Cited by 17 publications

(18 citation statements)

References 110 publications

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“…Many studies have shown that small proteins (≤100 amino acid residues) can be involved in important biological processes, including cell signaling, metabolism, and growth [21]. However, they are underrepresented in many genome annotations as they are notoriously hard to predict because of their small ORF size [21]. To validate these small peptides, we kept all ORF with at least 20 amino acids.…”

Section: Discussionmentioning

confidence: 99%

Proteotranscriptomics assisted gene annotation and spatial proteomics of Bombyx mori BmN4 cell line

2020

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Background The process of identifying all coding regions in a genome is crucial for any study at the level of molecular biology, ranging from single-gene cloning to genome-wide measurements using RNA-seq or mass spectrometry. While satisfactory annotation has been made feasible for well-studied model organisms through great efforts of big consortia, for most systems this kind of data is either absent or not adequately precise. Results Combining in-depth transcriptome sequencing and high resolution mass spectrometry, we here use proteotranscriptomics to improve gene annotation of protein-coding genes in the Bombyx mori cell line BmN4 which is an increasingly used tool for the analysis of piRNA biogenesis and function. Using this approach we provide the exact coding sequence and evidence for more than 6200 genes on the protein level. Furthermore using spatial proteomics, we establish the subcellular localization of thousands of these proteins. We show that our approach outperforms current Bombyx mori annotation attempts in terms of accuracy and coverage. Conclusions We show that proteotranscriptomics is an efficient, cost-effective and accurate approach to improve previous annotations or generate new gene models. As this technique is based on de-novo transcriptome assembly, it provides the possibility to study any species also in the absence of genome sequence information for which proteogenomics would be impossible.

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Section: Discussionmentioning

confidence: 99%

Proteotranscriptomics assisted gene annotation and spatial proteomics of Bombyx mori BmN4 cell line

2020

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“…A compounding issue in emerging new fields is the development of a consensus on nomenclature [11]. As the underlying genes are denoted either as small or short ORFs, with both terms being used interchangeably, their gene products are hence named either small open reading frame encoded peptides (SEPs).…”

Section: Statement Of Significancementioning

confidence: 99%

Bottom‐up and top‐down proteomic approaches for the identification, characterization, and quantification of the low molecular weight proteome with focus on short open reading frame‐encoded peptides

et al. 2021

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The recent discovery of alternative open reading frames creates a need for suitable analytical approaches to verify their translation and to characterize the corresponding gene products at the molecular level. As the analysis of small proteins within a background proteome by means of classical bottom-up proteomics is challenging, method development for the analysis of small open reading frame encoded peptides (SEPs) have become a focal point for research. Here, we highlight bottom-up and top-down proteomics approaches established for the analysis of SEPs in both pro-and eukaryotes. Major steps of analysis, including sample preparation and (small) proteome isolation, separation and mass spectrometry, data interpretation and quality control, quantification, the analysis of post-translational modifications, and exploration of functional aspects of the SEPs by means of proteomics technologies are described. These methods do not exclusively cover the analytics of SEPs but simultaneously include the low molecular weight proteome, and moreover, can also be used for the proteome-wide analysis of proteolytic processing events.

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“…The nomenclature and definition of the maximal size of the small proteins encoded by sORFs varies between 50 and 100 amino acids . Typically, peptides are distinguished from proteins by the shorter length of their chain.…”

Section: Introductionmentioning

confidence: 99%

Rapid Biophysical Characterization and NMR Spectroscopy Structural Analysis of Small Proteins from Bacteria and Archaea

et al. 2020

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Proteins encoded by small open reading frames (sORFs) have a widespread occurrence in diverse microorganisms and can be of high functional importance. However, due to annotation biases and their technically challenging direct detection, these small proteins have been overlooked for a long time and were only recently rediscovered. The currently rapidly growing number of such proteins requires efficient methods to investigate their structure–function relationship. Herein, a method is presented for fast determination of the conformational properties of small proteins. Their small size makes them perfectly amenable for solution‐state NMR spectroscopy. NMR spectroscopy can provide detailed information about their conformational states (folded, partially folded, and unstructured). In the context of the priority program on small proteins funded by the German research foundation (SPP2002), 27 small proteins from 9 different bacterial and archaeal organisms have been investigated. It is found that most of these small proteins are unstructured or partially folded. Bioinformatics tools predict that some of these unstructured proteins can potentially fold upon complex formation. A protocol for fast NMR spectroscopy structure elucidation is described for the small proteins that adopt a persistently folded structure by implementation of new NMR technologies, including automated resonance assignment and nonuniform sampling in combination with targeted acquisition.

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Ribosomal Peptides and Small Proteins on the Rise

Cited by 17 publications

References 110 publications

Proteotranscriptomics assisted gene annotation and spatial proteomics of Bombyx mori BmN4 cell line

Proteotranscriptomics assisted gene annotation and spatial proteomics of Bombyx mori BmN4 cell line

Bottom‐up and top‐down proteomic approaches for the identification, characterization, and quantification of the low molecular weight proteome with focus on short open reading frame‐encoded peptides

Rapid Biophysical Characterization and NMR Spectroscopy Structural Analysis of Small Proteins from Bacteria and Archaea

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