title: Cooperation between an intrinsically disordered region and a helical segment is required for ubiquitin-independent degradation by the proteasome.
abstract: The 26 S proteasomal complex, which is responsible for the bulk of protein degradation within the cell, recognizes its target substrates via covalently linked polyubiquitin moieties. However, a small but growing number of proteasomal substrates are degraded without a requirement for ubiquitinylation. One such substrate is the pyrimidine biosynthetic enzyme thymidylate synthase (EC 2.1.1.45), which catalyzes the synthesis of TMP and is the sole de novo source of TTP for DNA replication and repair. Previous work showed that intracellular proteolysis of human thymidylate synthase is directed by a degron at the polypeptide's N-terminal end, composed of an intrinsically disordered region (IDR) followed by a highly conserved amphipathic α-helix (hA). In the present report, we show that the hA helix does not function simply as an extension or scaffold for the IDR; rather, it provides a specific structural component that is necessary for degradation. Furthermore, its helical conformation is required for this function. We demonstrate that small domains from heterologous proteins can substitute for the IDR and the hA helix of human thymidylate synthase, indicating that the degradation-promoting function of these regions is not sequence-specific. The results, in general, indicate that cooperation between intrinsically disordered domains and α-helical segments is required for ubiquitin-independent degradation by the proteasome. There appears to be little sequence constraint on the ability of these regions to function as degron constituents. Rather, it is the overall conformation (or lack thereof) that is critical.
authors: Department of Biological Sciences and Center for Colon Cancer Research, University of South Carolina, Columbia, South Carolina 29208, USA.
authors: Sandra P
authors: SP
authors: Melo
authors: Karen W
authors: KW
authors: Barbour
authors: Franklin G
authors: FG
authors: Berger
mesh_terms: D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D051379:Mice; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism*; D017433:Protein Structure, Secondary; D059748:Proteolysis; D013940:Thymidylate Synthase / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology*
chemical_list: D025801:Ubiquitin; D013940:Thymidylate Synthase; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease