Methods in molecular biology
-
Post-translational modifications (PTMs) are one of the main contributors to the diversity of proteoforms in the proteomic landscape. In particular, protein phosphorylation represents an essential regulatory mechanism that plays a role in many biological processes. Protein kinases, the enzymes catalyzing this reaction, are key participants in metabolic and signaling pathways. ⋯ In this chapter, we demonstrate two text mining tools, RLIMS-P and eFIP, for the retrieval and extraction of kinase-substrate-site data and phosphorylation-dependent PPIs from the literature. These tools offer several advantages over a literature search in PubMed as their results are specific for phosphorylation. RLIMS-P and eFIP results can be sorted, organized, and viewed in multiple ways to answer relevant biological questions, and the protein mentions are linked to UniProt identifiers.
-
Recent advances in mass spectrometry based proteomic techniques and publicly available large proteomic repositories are being exploited to characterize the proteome of multiple organisms. While humongous amount of proteomic data is being acquired and analyzed, many biological questions still remain unanswered. Proteotypic peptides which uniquely represent target proteins or a protein isoform are used as an alternative strategy for protein identification in the field of immunological methods and targeted proteomic techniques. Using different computational approaches, resources and techniques used in the identification of proteotypic peptides of target proteins is discussed here.
-
Normal cellular functioning is maintained by macromolecular machines that control both core and specialized molecular tasks. These machines are in large part multi-subunit protein complexes that undergo regulation at multiple levels, from expression of requisite components to a vast array of post-translational modifications (PTMs). ⋯ Here we provide a framework for systematically studying these PTMs in the context of global protein-protein interaction networks. This analytical framework allows insight into which functions specific PTMs tend to cluster in, and furthermore which complexes either single or multiple PTM signaling pathways converge on.
-
Protein post-translational modification (PTM) is an essential cellular regulatory mechanism, and disruptions in PTM have been implicated in disease. PTMs are an active area of study in many fields, leading to a wealth of PTM information in the scientific literature. ⋯ This chapter describes the use of iPTMnet ( http://proteininformationresource.org/iPTMnet/ ), a resource that integrates PTM information from text mining, curated databases, and ontologies and provides visualization tools for exploring PTM networks, PTM crosstalk, and PTM conservation across species. We present several PTM-related queries and demonstrate how they can be addressed using iPTMnet.
-
Exon-skipping therapy is an emerging approach that uses synthetic DNA-like molecules called antisense oligonucleotides (AONs) to splice out frame-disrupting parts of mRNA, restore the reading frame, and produce truncated yet functional proteins. Multiple exon skipping utilizing a cocktail of AONs can theoretically treat 80-90% of patients with Duchenne muscular dystrophy (DMD). The success of multiple exon skipping by the systemic delivery of a cocktail of AONs called phosphorodiamidate morpholino oligomers (PMOs) in a DMD dog model has made a significant impact on the development of therapeutics for DMD, leading to clinical trials of PMO-based drugs. Here, we describe the systemic delivery of a cocktail of PMOs to skip multiple exons in dystrophic dogs and the evaluation of the efficacies and toxicity in vivo.