scholarly journals Quantifying homologous proteins and proteoforms

2017 ◽  
Author(s):  
Dmitry Malioutov ◽  
Tianchi Chen ◽  
Jacob Jaffe ◽  
Edoardo Airoldi ◽  
Steve Carr ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Alternative Splicing ◽  
Histone Modification ◽  
First Principles ◽  
High Accuracy ◽  
Biological Functions ◽  
Homologous Proteins ◽  
Bottom Up ◽  
Post Translational Modifications ◽  
Paralogous Genes

Many proteoforms – arising from alternative splicing, post-translational modifications (PTMs), or paralogous genes – have distinct biological functions, such as histone PTM proteoforms. However, their quantification by existing bottom-up mass–spectrometry (MS) methods is undermined by peptide-specific biases. To avoid these biases, we developed and implemented a first-principles model (HIquant) for quantifying proteoform stoichiometries. We characterized when MS data allow inferring proteoform stoichiometries by HIquant, derived an algorithm for optimal inference, and demonstrated experimentally high accuracy in quantifying fractional PTM occupancy without using external standards, even in the challenging case of the histone modification code. A HIquant server is implemented at: https://web.northeastern.edu/slavov/2014_HIquant/

scholarly journals VectorMOD: Method for Bottom-Up Proteomic Characterization of rAAV Capsid Post-Translational Modifications and Vector Impurities

2021 ◽  
Vol 12 ◽  
Author(s):  
Neil G. Rumachik ◽  
Stacy A. Malaker ◽  
Nicole K. Paulk
Keyword(s):  
Mass Spectrometry ◽  
Process Development ◽  
Cell Protein ◽  
Macromolecular Complex ◽  
Bottom Up ◽  
Post Translational Modifications ◽  
Simple Step ◽  
Product And Process ◽  
High Doses ◽  
And Control

Progress in recombinant AAV gene therapy product and process development has advanced our understanding of the basic biology of this critical delivery vector. The discovery of rAAV capsid post-translational modifications (PTMs) has spurred interest in the field for detailed rAAV-specific methods for vector lot characterization by mass spectrometry given the unique challenges presented by this viral macromolecular complex. Recent concerns regarding immunogenic responses to systemically administered rAAV at high doses has highlighted the need for investigators to catalog and track potentially immunogenic vector lot components including capsid PTMs and PTMs on host cell protein impurities. Here we present a simple step-by-step guide for academic rAAV laboratories and Chemistry, Manufacturing and Control (CMC) groups in industry to perform an in-house or outsourced bottom-up mass spectrometry workflow to characterize capsid PTMs and process impurities.

scholarly journals A Peptidoform Matching Strategy in Bottom-Up Proteomics for Studying Functions of Post-Translational Modifications

2021 ◽  
Author(s):  
Yansheng Liu
Keyword(s):  
Quantitative Analysis ◽  
Relevant Information ◽  
Previous Literature ◽  
Biological Functions ◽  
Biological Investigation ◽  
Bottom Up ◽  
Post Translational Modifications ◽  
Site Specific ◽  
Specific Approach ◽  
Matching Strategy

Protein translational modifications (PTMs) generate an enormous, but as yet undetermined, expansion of the expressed proteoforms. In this Viewpoint, we firstly differentiate the concepts of proteoform and peptidoform by reviewing and discussing previous literature. We show that the current PTM biological investigation and annotation largely follow a PTM site-specific rather than proteoform-specific approach. We further illustrate a potentially useful matching strategy in which a particular “modified peptidoform” is matched to the corresponding “unmodified peptidoform” as a reference for the quantitative analysis between samples and conditions. We suggest this strategy could provide directly relevant information for learning the PTM site-specific biological functions. Accordingly, we advocate for the wider use of the nomenclature “peptidoform” in the future bottom-up proteomic studies.

4. Proteins

2016 ◽  
Author(s):  
Aysha Divan ◽  
Janice A. Royds
Keyword(s):  
Alternative Splicing ◽  
Human Genome ◽  
The Body ◽  
Biological Functions ◽  
Protein Coding ◽  
Post Translational Modifications ◽  
Protein Coding Genes ◽  
Composition And Structure ◽  
A Cell ◽  
Structure Of Proteins

Biological functions require protein and the protein makeup of a cell determines its behaviour and identity. Proteins, therefore, are the most abundant molecules in the body except for water. The approximately 20,000 protein coding genes in the human genome can, by alternative splicing, multiple translation starts, and post-translational modifications, produce over 1,000,000 different proteins, collectively called ‘the proteome’. It is the size of the proteome and not the genome that defines the complexity of an organism. ‘Proteins’ describes the composition and structure of proteins and how they are studied. What information is required in order to understand how proteins work and what happens when this function is impaired in disease?

scholarly journals Proteomics: a powerful tool to study plant responses to biotic stress

Plant Methods ◽  
2019 ◽  
Vol 15 (1) ◽  
Author(s):  
Yahui Liu ◽  
Song Lu ◽  
Kefu Liu ◽  
Sheng Wang ◽  
Luqi Huang ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Biotic Stress ◽  
Resistance Mechanisms ◽  
Current Status ◽  
Biological Research ◽  
Plant Responses ◽  
Biological Functions ◽  
Comprehensive Understanding ◽  
Post Translational Modifications ◽  
Research Challenges

AbstractIn recent years, mass spectrometry-based proteomics has provided scientists with the tremendous capability to study plants more precisely than previously possible. Currently, proteomics has been transformed from an isolated field into a comprehensive tool for biological research that can be used to explain biological functions. Several studies have successfully used the power of proteomics as a discovery tool to uncover plant resistance mechanisms. There is growing evidence that indicates that the spatial proteome and post-translational modifications (PTMs) of proteins directly participate in the plant immune response. Therefore, understanding the subcellular localization and PTMs of proteins is crucial for a comprehensive understanding of plant responses to biotic stress. In this review, we discuss current approaches to plant proteomics that use mass spectrometry, with particular emphasis on the application of spatial proteomics and PTMs. The purpose of this paper is to investigate the current status of the field, discuss recent research challenges, and encourage the application of proteomics techniques to further research.

Mass Spectrometric (MS) Analysis of Proteins and Peptides

2020 ◽  
Vol 21 ◽  
Author(s):  
Madhuri Jayathirtha ◽  
Emmalyn J. Dupree ◽  
Zaen Manzoor ◽  
Brianna Larose ◽  
Zach Sechrist ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Alternative Splicing ◽  
Human Genome ◽  
Mass Spectrometric ◽  
Protein Isoforms ◽  
Gene Products ◽  
Post Translational Modifications ◽  
The Past ◽  
Transient Interactions ◽  
Proteins And Peptides

: The human genome is sequenced and is comprised of~30,000 genes, making humans just a little bit more complicated than worms or flies. However, complexity of humans is given by proteins that these genes code for, because one gene can produce many proteins mostly through alternative splicing and tissue-dependent expression of particular proteins. In addition, post-translational modifications (PTMs) in proteins greatly increase the number of gene products or protein isoforms. Furthermore, stable and transient interactions between proteins, protein isoforms/proteoforms and PTM-ed proteins (proteinprotein interactions, PPI) adds yet another level of complexity in humans and other organisms. In the past, all of these proteins were analyzed one at the time. Currently, they are analyzed by a less tedious method: mass spectrometry (MS) for two reasons: 1) because of the complexity of proteins, protein PTMs and PPIs and 2) because MS is the only method that can keep up with such a complex array of features. Here, we discuss the applications of mass spectrometry in protein analysis.

scholarly journals Accelerating the field of epigenetic histone modification through mass spectrometry-based approaches

2020 ◽  
pp. mcp.R120.002257
Author(s):  
Congcong Lu ◽  
Mariel Coradin ◽  
Elizabeth G Porter ◽  
Benjamin A Garcia
Keyword(s):  
Mass Spectrometry ◽  
Molecular Biology ◽  
Histone Modification ◽  
High Throughput ◽  
Epigenetic Regulation ◽  
Associated Factors ◽  
Chemical Biology ◽  
Post Translational Modifications ◽  
Histone Ptms ◽  
Chromatin Biology

Histone post-translational modifications (PTMs) are one of the main mechanisms of epigenetic regulation. Dysregulation of histone PTMs leads to many human diseases, such as cancer. Due to its high-throughput, accuracy, and flexibility, mass spectrometry (MS) has emerged as a powerful tool in the epigenetic histone modification field, allowing the comprehensive and unbiased analysis of histone PTMs and chromatin-associated factors. Coupled with various techniques from molecular biology, biochemistry, chemical biology and biophysics, MS has been employed to characterize distinct aspects of histone PTMs in the epigenetic regulation of chromatin functions. In this review we will describe advancements in the field of MS that have facilitated the analysis of histone PTMs and chromatin biology.  

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