Proteomic insights into ubiquitin and ubiquitin-like proteins

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The dynamic and specific modification of cellular proteins by members of the ubiquitin protein family is a vital regulatory mechanism that lies at the heart of almost all biological processes. Because of both their pervasive and complex nature, these regulatory pathways have been the target of many recent proteomic studies. Such works have provided numerous insights. Through the use of various mass spectrometry techniques, affinity purification methods, and/or chemical probes, large lists have begun to be compiled for the multitude of substrates, interacting partners, and enzymatic components of these regulatory circuits. Furthermore, similar tools have provided many insights into functional aspects such as their mechanisms of substrate specificity and enzymatic activity. This review provides a summary of these recent proteomic works, along with comments on future directions of the field.

Introduction

Post-translational modifications (PTMs) of cellular proteins act to rapidly and precisely modulate a diverse array of biological processes. Many of the most important PTM regulatory switches belong to the ubiquitin (Ub) protein family, including Ub, SUMO (small ubiquitin-like modifier), and numerous other Ub-like proteins (Ubls). Given their functional and biochemical diversity, these pathways are particularly well suited for proteomic-type analyses. This review focuses on recent works that have successfully used proteomic tools to elucidate the protein substrates, the interacting partners and the enzymatic components that comprise the Ub and Ubl-systems.

Section snippets

Proteomic approaches for identifying the components of Ub- and Ubl-systems

The biochemical mechanisms and functions of Ub 1., 2., 3., 4., SUMO [5] and other Ubl modifications [6] have been thoroughly reviewed elsewhere, with the fundamental component common to each of these pathways being the formation of a covalent bond between the C-terminal glycine residue of Ub/Ubl and a ɛ-amino group of a substrate lysine [1]. To further understand the biological consequences of covalent adducts generated in this manner, a large body of work has focused on using MS-based

Using proteomics to unravel the function of Ub/Ubl pathways

An important step in uncovering the functional relevance of protein ubiquitination (or Ubl-modification) is to identify the actual sites of conjugation on substrates. Once Ub/Ubl-modification sites are identified, site-directed mutagenesis can be performed on these specific lysine residues to test the biological consequences of the PTM. Towards this end, MS-based approaches have been particularly useful. Unlike non-protein PTMs such as phosphorylation and methylation, the identification of

Closing comments and future directions

As summarized in Figure 2 for Ub, numerous aspects of Ub- and Ubl-pathways have been targeted by proteomic studies in recent years. These works have combined to greatly enhance our understanding of the identity and function of the substrates, binding partners, and effector enzymes of these systems. This has been aided tremendously by the fact that MS provides a means of identifying polypeptides without any prior knowledge of their identity or function, thereby allowing for the discovery of

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

Acknowledgements

This work was supported by National Institutes of Health grants HG00041 and GM67945.

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