Reviews & Perspectives
An Expanding Repertoire of Protein Acylations
In Brief In this work, we give a general overview of the 12 main protein acylations, also including novel acylations, such as benzoylation and 2-hydroxyisobutyrylation. We summarize the recent advances in protein acylation, mainly focus on their substrates, enzymes, biological functions, and novel detecting methods and related diseases, especially in cancer. We believe that the review will provide an unprecedented and comprehensive view of protein acylations and bring important reference significance for future research.Subcellular Transcriptomics and Proteomics: A Comparative Methods Review
In Brief The interior of the cell is molecularly crowded. Its compartmentalization within organelles enables the regulation of biochemical processes and allows multifunctionality of proteins and RNAs. Subcellular information can thus give insights into the function of these biomolecules. Multiple techniques to measure such information have been established, with ever-increasing throughput and sensitivity. These techniques are covered in this review, and demonstrating their application is providing valuable insights into cellular biology, such as aiding our understanding of single-cell heterogeneity and posttranslational modifications.Reflections on the HUPO Human Proteome Project, the Flagship Project of the Human Proteome Organization, at 10 Years
In Brief Starting from several organ-oriented projects, HUPO in 2010 launched the Human Proteome Project to identify and characterize the protein parts list and integrate proteomics into multiomics research. Key steps were partnerships with neXtProt, PRIDE, PeptideAtlas, Human Protein Atlas, and instrument makers; global engagement of researchers; creation of ProteomeXchange; adoption of HPP Guidelines for Interpretation of MS Data and SRMAtlas for proteotypic peptides; annual metrics of finding “missing proteins” and functionally annotating proteins; and initiatives for early career scientists.Data Management of Sensitive Human Proteomics Data: Current Practices, Recommendations, and Perspectives for the Future
In Brief Availability of proteomics data in the public domain has become the norm, as it has been the case in genomics and transcriptomics for many years. Analogously to sequencing data, there are increasing ethical issues and legal requirements related to sensitive human clinical proteomics data. We review the current state of the art and make concrete recommendations to address these issues in the proteomics field, which are summarized in four different areas.Neuroproteomics of the Synapse: Subcellular Quantification of Protein Networks and Signaling Dynamics
In Brief Advancements in MS-based proteomics have increased the study of synaptic proteins using neuroproteomics. The development of proximity, genetic labeling and bio-orthogonal amino acid labeling approaches now allow for the study of synaptic protein–protein interactions and protein signaling dynamics. In this review, we highlight studies from the last 5 years, with a focus on synapse structure, composition, functioning, or signaling and finally discuss some recent developments that could further advance the field of neuroproteomics.Decoding Post-Translational Modification Crosstalk With Proteomics
In Brief We provide an overview of current experimental and computational proteomic methods, as well as a perspective on emerging technologies to study PTM crosstalk.Recent Advances in Software Tools for More Generic and Precise Intact Glycopeptide Analysis
In Brief This article provides a systematic review of the most recent MS-based strategies and corresponding software tools for the analysis of intact glycopeptides, particularly intact N-glycopeptides, reported in the last decade, including the process of identifying N-glycopeptides from MS data, the existing methods of MS data acquisition and interpretation, the quality control methods, the display of results, and the software applications.Recent Advances in Analytical Approaches for Glycan and Glycopeptide Quantitation
In Brief Recent years have seen an explosion in novel strategies for quantitative glycomics and glycoproteomics. Whether through metabolic incorporation of stable isotopes, deposition of custom isotopic labels, or high-throughput isobaric chemical tags, these numerous novel strategies provide ease of access to glycomic and glycoproteomic investigation. This review highlights the recent innovations in labeling methods, label-free strategies, acquisition modes, and bioinformatic tools for glycan and glycopeptide quantitation, while providing critical evaluations and technical considerations to enable effective analysis.Developments in Mass Spectrometry for Glycosaminoglycan Analysis: A Review
In Brief Glycosaminoglycans (GAGs) participate in a variety of biological functions and have a multitude of medicinal properties. Due to their non template driven biosynthesis, GAGs are produced as nonuniform complex mixtures. Mass spectrometry paired with on-line separation techniques has been utilized to determine the composition of these complex mixtures. Advances in tandem mass spectrometry have also made determining sequence information such as sulfation location and C-5 epimerization possible. This review covers recent developments in the analysis of GAGs using mass spectrometry.Methods for Enrichment and Assignment of N-Acetylglucosamine Modification Sites
In Brief This review article summarizes methods for O-GlcNAc enrichment and different mass spectrometric approaches for acquiring data on modified peptides and describes software strategies for analyzing data, including the challenges of reliably identifying modification sites and differentiating between other potential HexNAc modifications. It then presents a new dataset to exemplify what is currently achievable.A Pragmatic Guide to Enrichment Strategies for Mass Spectrometry–Based Glycoproteomics
In Brief Interest in mass spectrometry–based glycoproteomics analysis is increasing because of recent advances in instrumentation and data analysis tools. Such studies can provide a wealth of information across a wide spectrum of glycan classes and biological systems. However, many studies require the choice of an enrichment strategy for glycosylated species prior to analysis to obtain the maximum amount of analytical information. Here, common enrichment strategies are reviewed with strengths and weaknesses, and the practical considerations for various methods are discussed.Accelerating the Field of Epigenetic Histone Modification Through Mass Spectrometry–Based Approaches
In Brief Histone post-translational modifications play essential roles in the epigenetic regulation of chromatin-related functions. Because of its high throughput, accuracy, and flexibility, mass spectrometry has emerged as a powerful tool in the epigenetic field. In this review, we describe the contributions of mass spectrometry–based proteomics in combination with distinct labeling strategies and various biological techniques to understand the roles of histone post-translational modifications and how they regulate chromatin function.Organellar Maps Through Proteomic Profiling – A Conceptual Guide
Protein subcellular localization is highly regulated and critical for protein function. Spatial proteomics aims at capturing the localization dynamics of all proteins expressed in a given cell type. Among different approaches, organellar mapping through proteomic profiling stands out as the only method capable of determining the subcellular localizations of thousands of proteins in a single experiment. Importantly, it can also detect movements of proteins between subcellular compartments, providing an unbiased systems analysis tool for investigating physiological and pathological cellular processes.Peptide-based Interaction Proteomics
Protein-protein interactions that are mediated by short linear motifs (SLiMs) in intrinsically disordered regions (IDRs) of proteins are notoriously difficult to study. Recently, pull-downs with synthetic peptides in combination with quantitative mass spectrometry emerged as a powerful screening approach. Here, we briefly highlight the relevance of SLiMs for protein-protein interactions, outline existing screening technologies, discuss unique advantages of peptide-based interaction screens, and provide practical suggestions for setting up such peptide-based screens.Proximity Dependent Biotinylation: Key Enzymes and Adaptation to Proteomics Approaches
Proximity-dependent biotinylation approaches such as BioID and APEX overcome classical limitations of biochemical purification and have gained widespread use in recent years for revealing cellular neighborhoods. Here we focus on the structural diversity and mechanisms of the two classes of enzymes, biotin protein ligases and peroxidases, and discuss current and emerging applications of these enzymes for proximity dependent biotinylation. We provide guidelines for enzyme selection and experimental design for performing and interpreting proximity-dependent biotinylation experiments.Next-generation Interactomics: Considerations for the Use of Co-elution to Measure Protein Interaction Networks
Interactome studies are necessary to understand cellular processes and co-elution methods are well suited for the simultaneous and global exploration of the interactome, as well as the assessment of biological perturbations of the network. These methods rely on the fundamental idea that proteins from the same complex migrate together during fractionation. We review the different separation techniques along with the quantification and bioinformatic approaches used for co-elution methods and provide design considerations to choose between them.
