5LOZ

STRUCTURE OF YEAST ENT1 ENTH DOMAIN

Summary for 5LOZ

• Entry DOI: 10.2210/pdb5loz/pdb
• Related: 1EDU 1EYH 1HOA
• Descriptor: Epsin-1 (2 entities in total)
• Functional Keywords: alpha-alpha superhelix, ubiquitin receptor, endocytosis adaptor, ubiquitin, clathrin, lipid, eps15, ubiquitin-binding domain
• Biological source: Saccharomyces cerevisiae (Baker's yeast)
• Cellular location: Cytoplasm: Q12518
• Total number of polymer chains: 1
• Total formula weight: 15981.05

Authors

Tanner, N.,Prag, G. (deposition date: 2016-08-11, release date: 2016-10-05, Last modification date: 2024-05-08)

Primary citation

Levin-Kravets, O.,Tanner, N.,Shohat, N.,Attali, I.,Keren-Kaplan, T.,Shusterman, A.,Artzi, S.,Varvak, A.,Reshef, Y.,Shi, X.,Zucker, O.,Baram, T.,Katina, C.,Pilzer, I.,Ben-Aroya, S.,Prag, G.
A bacterial genetic selection system for ubiquitylation cascade discovery.
Nat.Methods, 13:945-952, 2016

PubMed Abstract: About one-third of the eukaryotic proteome undergoes ubiquitylation, but the enzymatic cascades leading to substrate modification are largely unknown. We present a genetic selection tool that utilizes Escherichia coli, which lack deubiquitylases, to identify interactions along ubiquitylation cascades. Coexpression of split antibiotic resistance protein tethered to ubiquitin and ubiquitylation target together with a functional ubiquitylation apparatus results in a covalent assembly of the resistance protein, giving rise to bacterial growth on selective media. We applied the selection system to uncover an E3 ligase from the pathogenic bacteria EHEC and to identify the epsin ENTH domain as an ultraweak ubiquitin-binding domain. The latter was complemented with a structure-function analysis of the ENTH-ubiquitin interface. We also constructed and screened a yeast fusion library, discovering Sem1 as a novel ubiquitylation substrate of Rsp5 E3 ligase. Collectively, our selection system provides a robust high-throughput approach for genetic studies of ubiquitylation cascades and for small-molecule modulator screening.

PubMed: 27694912
DOI: 10.1038/nmeth.4003

Experimental method

X-RAY DIFFRACTION (1.95 Å)