title: Advances in purification and separation of posttranslationally modified proteins.
abstract: Posttranslational modifications (PTMs) of proteins represent fascinating extensions of the dynamic complexity of living cells' proteomes. The results of enzymatically catalyzed or spontaneous chemical reactions, PTMs form a fourth tier in the gene - transcript - protein cascade, and contribute not only to proteins' biological functions, but also to challenges in their analysis. There have been tremendous advances in proteomics during the last decade. Identification and mapping of PTMs in proteins have improved dramatically, mainly due to constant increases in the sensitivity, speed, accuracy and resolution of mass spectrometry (MS). However, it is also becoming increasingly evident that simple gel-free shotgun MS profiling is unlikely to suffice for comprehensive detection and characterization of proteins and/or protein modifications present in low amounts. Here, we review current approaches for enriching and separating posttranslationally modified proteins, and their MS-independent detection. First, we discuss general approaches for proteome separation, fractionation and enrichment. We then consider the commonest forms of PTMs (phosphorylation, glycosylation and glycation, lipidation, methylation, acetylation, deamidation, ubiquitination and various redox modifications), and the best available methods for detecting and purifying proteins carrying these PTMs. This article is part of a Special Issue entitled: Posttranslational Protein modifications in biology and Medicine.
authors.affiliation: Department of Molecular Biology and Radiobiology, Mendel University in Brno & CEITEC - Central European Institute of Technology, Mendel University in Brno, Zemědělská 1, CZ-613 00 Brno, Czech Republic. Electronic address: martincerny83@gmail.com.
authors.forename: Martin
authors.initials: M
authors.lastname: Černý
authors.forename: Jan
authors.initials: J
authors.lastname: Skalák
authors.forename: Hana
authors.initials: H
authors.lastname: Cerna
authors.forename: Břetislav
authors.initials: B
authors.lastname: Brzobohatý
mesh_terms: D000818:Animals; D006801:Humans; D011499:Protein Processing, Post-Translational; D020543:Proteome / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*
chemical_list: D020543:Proteome
keywords: PTM; PTM detection; Posttranslational modification; Protein enrichment; Proteome; Top-down proteomics