N-terminomics - its past and recent advancements.

Abstract N-terminomics is a rapidly evolving branch of proteomics that encompasses the study of protein N-terminal sequence. A proteome-wide collection of such sequences has been widely used to understand the proteolytic cascades and in annotating the genome. Over the last two decades, various N-terminomic strategies have been developed for achieving high sensitivity, greater depth of coverage, and high-throughputness. We, in this review, cover how the field of N-terminomics has evolved to date, including discussion on various sample preparation and N-terminal peptide enrichment strategies. We also compare different N-terminomic methods and highlight their relative benefits and shortcomings in their implementation. In addition, an overview of the currently available bioinformatics tools and data analysis pipelines for the annotation of N-terminomic datasets is also included. Significance It has been recognized that proteins undergo several post-translational modifications (PTM), and a number of perturbed biological pathways are directly associated with modifications at the terminal sites of a protein. In this regard, N-terminomics can be applied to generate a proteome-wide landscape of mature N-terminal sequences, annotate their source of generation, and recognize their significance in the biological pathways. Besides, a system-wide study can be used to study complicated proteolytic machinery and protease cleavage patterns for potential therapeutic targets. Moreover, due to unprecedented improvements in the analytical methods and mass spectrometry instrumentation in recent times, the N-terminomic methodologies now offers an unparalleled ability to study proteoforms and their implications in clinical conditions. Such approaches can further be applied for the detection of low abundant proteoforms, annotation of non-canonical protein coding sites, identification of candidate disease biomarkers, and, last but not least, the discovery of novel drug targets.