The Eukaryotic Linear Motif resource for
Functional Sites in Proteins
Functional site class:
C-terminal cyclic imide degron motif binding the TBD domain of Cereblon
Functional site description:
Cereblon (CRBN) is a substrate receptor of the E3 cullin 4–RING ligase (CRL4) complex and is a direct target of thalidomide and other immunomodulatory drugs (IMiDs). The C-terminal thalidomide-binding domain acts as a binding site for IMiDs but the natural degrons are C-terminal cyclic imides. Interaction with IMiDs modulates the substrate specificity of cereblon and enables the binding of so-called neo-substrate proteins to the ligase complex for ubiquitination and subsequent proteasomal degradation: this has many therapeutical implications, as in the case of hematologic malignancies. Recognition of the natural degrons like aspartimide and aminoglutarimide residues helps in removing malfunctional and harmful fragments. The thalidomide-binding domain of cereblon is highly conserved in both eukaryotes and prokaryotes and suggests that it might bind a common ligand and have a universal cellular function. Cereblon-binding IMiDs are potential teratogens but not all species are susceptible to the same extent.
ELM Description:
CRBN recognizes C-terminal cyclic imides, specifically aspartimide (amino succinimide) and aminoglutarimide residues as its natural degrons. These are formed as a result of spontaneous chain breaks after an attack of an asparagine or glutamine side chain amide on the adjacent peptide bond. The crystal structure of the APP (QMQcN peptide) - MsCI4 8BC6) complex shows that only the C-terminal residue of the cyclic imide (here, succinimide ring) is directly slotted into a tri- tryptophan cage (W79, W85, W99 in MsCI4) present in the thalidomide-binding domain (TBD) while the three other residues of the peptide are protruding outside and make hydrogen bonds between peptide backbone oxygens and strictly conserved CRBN residues. The binding affinities increase with peptide length. As in the case of a 4-mer, the Cterm-1 and Cterm-2 positions form hydrogen bonds with N50/(hN351) and W99/(hW400), respectively, while the Cterm-3 residue is forming a second hydrogen bond with the N50/N351 side chain. While comparing the recognition modes of peptides with a C-terminal aspartimide or aminoglutarimide residue, they are essentially the same and bind with the same efficiency.
Studies have shown that CRBN specifically recognizes only the cyclic modifications at the C-terminus, but not the N-terminus (Heim,2022). Most of the IMiDs like thalidomide, lenalidomide and pomalidomide uses the same mechanism for CRBN binding where a glutarimide moiety is inserted into the aromatic cage and phthalimide carbonyl groups form a hydrogen bond with a conserved asparagine residue of the protein (Fischer,2014).

Because this C-terminal degron is caused by internal protein damage at any N or Q, these internal instances cannot be effectively represented in the ELM database.
Pattern: ...[NQ]$
Pattern Probability: 0.0001614
Present in taxons: Bacteria Eukaryota
Not represented in taxon: Fungi
Interaction Domain:
CULT domain (IPR034750) The cereblon protein, originally identified in a screen for mutations causing mild mental retardation, is a major target of thalidomide and its derivatives, and is responsible for the teratogenic effects of the drug (Stochiometry: 1 : 1)
o Abstract
The drug thalidomide was used in the 1950s-1960s as a sedative for pregnant women suffering acute morning sickness. Unfortunately, it proved strongly teratogenic, causing developmental abnormalities. For many years the molecular mechanism of the drug was unknown, although it had become clear that the drug might be a useful cancer treatment. In 2010, Ito et al. reported that thalidomide bound to the protein cereblon (CRBN), also demonstrating that the protein is an E3 ligase (Ito,2010), thus paving the way to understanding the molecular and cellular processes modulated by thalidomide.

Cereblon (CRBN) is a ubiquitously expressed protein that acts as a substrate receptor of the Cullin-4 E3 ubiquitin ligase complex, CUL4-RBX1-DDB1 (CRL4). The CRBN gene was initially identified in connection with memory and learning. Mutations in this gene cause a mild type of recessive nonsyndromic mental retardation (Higgins,2004). Later, it was reported to be involved in various biological activities such as the regulation of ion transport, modulation of the AMPK signalling pathway, and cell metabolism. Thalidomide and its analogues are termed Immunomodulatory imide drugs (IMiDs) as they adjust immune responses. It was discovered that IMiDs enable certain proteins to bind CRBN and be destroyed by CUL4-CRBN. These proteins are termed neo-substrates and include IRF4, IKZF1 and IKZF3, MYC and SALL4 (Costacurta,2021).

Repurposing its interactome by interacting with IMiDs like thalidomide and its analogues or proteolysis targeting chimeras (PROTACs) helps in the control of tumour growth and angiogenesis in adults (Ito,2021). By contrast, thalidomide is teratogenic for the developing foetus (Asatsuma-Okumura,2020).
The more recently identified cellular ligands also share the same binding groove as Thalidomide and its derivatives (Heim,2022). The C-terminal cyclic amides aspartimide and aminoglutarimide residues are the first natural degron motifs that are recognized by CRBN with high specificity. This post-translational modification is formed as a result of the intramolecular cyclization of glutamine or asparagine residues and concomitant cleavage of the peptide backbone. This reaction results in a fragmented protein which may have lost functionality, especially for any multidomain proteins where the modules interact with each other. Cyclic amides can arise spontaneously and are likely to accumulate in ageing proteins, and their detection acts as a mechanism for the efficient removal of malfunctional and damaged proteins. Moreover, these cyclic imides readily interact with free amino groups and can form additional unwanted cross-links that are frequently seen in many age-related diseases (Friedrich,2019). The best characterized cyclic imides are from Beta-crystallin and Amyloid-beta precursor proteins (Heim,2022; Ichikawa,2022). The CULT domain (IPR034750) of CRBN is present in bacteria as well as eukaryotes (though absent from fungi), suggesting that the removal of cyclic C-termini is also important in prokaryotes.

The C-terminal thalidomide-binding domain (TBD), also known as the CULT domain (Cereblon domain of Unknown activity, binding cellular Ligands and Thalidomide), acts as a binding site for the cellular degrons and IMiDs. It is highly conserved across species with very few residue substitutions and contains three strictly conserved tryptophan residues (tri-tryptophan pocket) that are important for ligand binding. In human cereblon (Q96SW2), Valine 388 is an important residue as the substitution of this residue to a smaller (V388A) or larger (V388I) side chain impaired neo-substrate binding and causes the teratogenic effect of thalidomide differently. Thalidomide is teratogenic in humans, zebrafish, and chicks but not in rodents (Heim,2023). Degradation of the SALL4 neosubstrate is likely to be particularly important for the teratogenicity of IMiDs (Donovan,2018).

Immunomodulatory drugs (IMiDs) or cereblon (CRBN) E3 ligase modulatory drugs (CELMoDs) are molecular glues. They recruit neo-substrate proteins to CRBN for ubiquitination and subsequent proteasomal degradation. The glutarimide ring of the cereblon binding moiety is slotted into the cage while the remaining molecules protrude out and act as a binding site for various neo-substrates with its specificity fine-tuned by the protruding moiety (Boichenko,2018). Several structures of neo-substrates in complex with CRBN and IMiDs are available (5HXB; 5FQD; 7U8F; 6H0F; 7BQV). These all show substantial interactions of globular domains; for example, C2H2 Zinc fingers insert into the CRBN interaction groove in the case of SALL4 and Ikaros transcription factors.
o 9 selected references:

o 7 GO-Terms:

o No Instance for DEG_CRBN_cyclicCter_1
Please cite: The Eukaryotic Linear Motif resource: 2022 release. (PMID:34718738)

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