@inbook{172fe77af92d41a080b9cebba92b4344,
title = "Identification of SUMO-interacting proteins by yeast two-hybrid analysis",
abstract = "This chapter will discuss various adaptations of the yeast two-hybrid method for analyzing protein interactions that can be used to identify small ubiquitin-related modifier (SUMO) interacting proteins and to determine the nature of the SUMO-protein interactions that occur. SUMO binds to a protein in two different ways: covalently and noncovalently. In a covalent interaction an isopeptide bond forms between the glycine residue at the C terminus of the mature SUMO and a lysine side-chain on the substrate protein. Alternatively, SUMO can interact noncovalently with another protein, usually via insertion of a β strand from a substrate SUMO-interacting motif (SIM) into a hydrophobic groove next to the SUMO β2 strand. By mutating either the C-terminal diglycine motif or amino acids within the β2 strand of SUMO, these respective interactions can be abolished. The expression of the two-hybrid SUMO constructs with either of these mutations can help distinguish the type of interaction that occurs between a SUMO and a given protein. Sumoylation can be verified by independent methods, such as a SUMO mobility shift assay. Finally, the chapter will compare the two-hybrid approach with mass spectrometric analysis as a means of identifying SUMO-interacting proteins.",
keywords = "Desumoylating enzymes, SIM (SUMO-interacting motif), SUMO, SUMO proteases, Two-hybrid analysis",
author = "Kroetz, {Mary B.} and Mark Hochstrasser",
year = "2009",
doi = "10.1007/978-1-59745-566-4_7",
language = "English (US)",
isbn = "9781934115800",
series = "Methods in Molecular Biology",
pages = "107--120",
editor = "Ulrich, {Helle D.}",
booktitle = "SUMO Protocols",
}