Ubiquitin ligase

File:Ubiquitylation.png

The ubiquitin ligase is referred to as an E3, and operates in conjunction with an E1 ubiquitin-activating enzyme and an E2 ubiquitin-conjugating enzyme. There is one major E1 enzyme, shared by all ubiquitin ligases, that uses ATP to activate ubiquitin for conjugation and transfers it to an E2 enzyme. The E2 enzyme interacts with a specific E3 partner and transfers the ubiquitin to the target protein. The E3, which may be a multi-protein complex, is, in general, responsible for targeting ubiquitination to specific substrate proteins.{{citation needed|date=March 2015}}

The ubiquitylation reaction proceeds in three or four steps depending on the mechanism of action of the E3 ubiquitin ligase. In the conserved first step, an E1 cysteine residue attacks the ATP-activated C-terminal glycine on ubiquitin, resulting in a thioester Ub-S-E1 complex. The energy from ATP and diphosphate hydrolysis drives the formation of this reactive thioester, and subsequent steps are thermoneutral. Next, a transthiolation reaction occurs, in which an E2 cysteine residue attacks and replaces the E1. HECT domain type E3 ligases will have one more transthiolation reaction to transfer the ubiquitin molecule onto the E3, whereas the much more common RING finger domain type ligases transfer ubiquitin directly from E2 to the substrate.{{cite journal | vauthors = Metzger MB, Hristova VA, Weissman AM | title = HECT and RING finger families of E3 ubiquitin ligases at a glance | journal = Journal of Cell Science | volume = 125 | issue = Pt 3 | pages = 531–7 | date = February 2012 | pmid = 22389392 | pmc = 3381717 | doi = 10.1242/jcs.091777 }} The final step in the first ubiquitylation event is an attack from the target protein lysine amine group, which will remove the cysteine, and form a stable isopeptide bond.{{cite book |last=Walsh |first=Christopher |title=Posttranslational Modification of Proteins: Expanding Nature's Inventory |location=Englewood, CO |publisher=Roberts |year=2006 |isbn=978-0-9747077-3-0}}{{page needed|date=March 2015}} One notable exception to this is p21 protein, which appears to be ubiquitylated using its N-terminal amine, thus forming a peptide bond with ubiquitin.{{cite journal | vauthors = Bloom J, Amador V, Bartolini F, DeMartino G, Pagano M | title = Proteasome-mediated degradation of p21 via N-terminal ubiquitinylation | journal = Cell | volume = 115 | issue = 1 | pages = 71–82 | date = October 2003 | pmid = 14532004 | doi = 10.1016/S0092-8674(03)00755-4 | doi-access = free }}

Humans have an estimated 500-1000 E3 ligases, which impart substrate specificity onto the E1 and E2.{{cite journal | vauthors = Nakayama KI, Nakayama K | title = Ubiquitin ligases: cell-cycle control and cancer | journal = Nature Reviews. Cancer | volume = 6 | issue = 5 | pages = 369–81 | date = May 2006 | pmid = 16633365 | doi = 10.1038/nrc1881 | s2cid = 19594293 }} The E3 ligases are classified into four families: HECT, RING-finger, U-box, and PHD-finger. The RING-finger E3 ligases are the largest family and contain ligases such as the anaphase-promoting complex (APC) and the SCF complex (Skp1-Cullin-F-box protein complex). SCF complexes consist of four proteins: Rbx1, Cul1, Skp1, which are invariant among SCF complexes, and an F-box protein, which varies. Around 70 human F-box proteins have been identified.{{cite journal | vauthors = Jin J, Cardozo T, Lovering RC, Elledge SJ, Pagano M, Harper JW | title = Systematic analysis and nomenclature of mammalian F-box proteins | journal = Genes & Development | volume = 18 | issue = 21 | pages = 2573–80 | date = November 2004 | pmid = 15520277 | pmc = 525538 | doi = 10.1101/gad.1255304 }} F-box proteins contain an F-box, which binds the rest of the SCF complex, and a substrate binding domain, which gives the E3 its substrate specificity.

File:Ubiquitin Lysines.png

Ubiquitin signaling relies on the diversity of ubiquitin tags for the specificity of its message. A protein can be tagged with a single ubiquitin molecule (monoubiquitylation), or variety of different chains of ubiquitin molecules (polyubiquitylation).{{cite journal | vauthors = Behrends C, Harper JW | title = Constructing and decoding unconventional ubiquitin chains | journal = Nature Structural & Molecular Biology | volume = 18 | issue = 5 | pages = 520–8 | date = May 2011 | pmid = 21540891 | doi = 10.1038/nsmb.2066 | s2cid = 19237120 }} E3 ubiquitin ligases catalyze polyubiquitination events much in the same way as the single ubiquitylation mechanism, using instead a lysine residue from a ubiquitin molecule currently attached to substrate protein to attack the C-terminus of a new ubiquitin molecule. For example, a common 4-ubiquitin tag, linked through the lysine at position 48 (K48) recruits the tagged protein to the proteasome, and subsequent degradation. However, all seven of the ubiquitin lysine residues (K6, K11, K27, K29, K33, K48, and K63), as well as the N-terminal methionine are used in chains in vivo.

Monoubiquitination has been linked to membrane protein endocytosis pathways. For example, phosphorylation of the Tyrosine at position 1045 in the Epidermal Growth Factor Receptor (EGFR) can recruit the RING type E3 ligase c-Cbl, via an SH2 domain. C-Cbl monoubiquitylates EGFR, signaling for its internalization and trafficking to the lysosome.{{cite journal | vauthors = Bonifacino JS, Traub LM | title = Signals for sorting of transmembrane proteins to endosomes and lysosomes | journal = Annual Review of Biochemistry | volume = 72 | pages = 395–447 | year = 2003 | pmid = 12651740 | doi = 10.1146/annurev.biochem.72.121801.161800 }}

Monoubiquitination also can regulate cytosolic protein localization. For example, the E3 ligase MDM2 ubiquitylates p53 either for degradation (K48 polyubiquitin chain), or for nuclear export (monoubiquitylation). These events occur in a concentration dependent fashion, suggesting that modulating E3 ligase concentration is a cellular regulatory strategy for controlling protein homeostasis and localization.{{cite journal | vauthors = Li M, Brooks CL, Wu-Baer F, Chen D, Baer R, Gu W | title = Mono- versus polyubiquitination: differential control of p53 fate by Mdm2 | journal = Science | volume = 302 | issue = 5652 | pages = 1972–5 | date = December 2003 | pmid = 14671306 | doi = 10.1126/science.1091362 | bibcode = 2003Sci...302.1972L | s2cid = 43124248 }}

Ubiquitin ligases are the final, and potentially the most important determinant of substrate specificity in ubiquitination of proteins.{{cite journal | vauthors = Zheng N, Shabek N | title = Ubiquitin Ligases: Structure, Function, and Regulation | journal = Annual Review of Biochemistry | volume = 86 | issue = 1 | pages = 129–157 | date = June 2017 | pmid = 28375744 | doi = 10.1146/annurev-biochem-060815-014922 }} The ligases must simultaneously distinguish their protein substrate from thousands of other proteins in the cell, and from other (ubiquitination-inactive) forms of the same protein. This can be achieved by different mechanisms, most of which involve recognition of degrons: specific short amino acid sequences or chemical motifs on the substrate.{{cite journal | vauthors = Ravid T, Hochstrasser M | title = Diversity of degradation signals in the ubiquitin-proteasome system | journal = Nature Reviews Molecular Cell Biology | volume = 9 | issue = 9 | pages = 679–90 | date = September 2008 | pmid = 18698327 | pmc = 2606094 | doi = 10.1038/nrm2468 }}

Proteolytic cleavage can lead to exposure of residues at the N-terminus of a protein. According to the N-end rule, different N-terminal amino acids (or N-degrons) are recognized to a different extent by their appropriate ubiquitin ligase (N-recognin), influencing the half-life of the protein.{{cite journal | vauthors = Sriram SM, Kim BY, Kwon YT | title = The N-end rule pathway: emerging functions and molecular principles of substrate recognition | journal = Nature Reviews Molecular Cell Biology | volume = 12 | issue = 11 | pages = 735–47 | date = October 2011 | pmid = 22016057 | doi = 10.1038/nrm3217 | s2cid = 10555455 }} For instance, positively charged (Arg, Lys, His) and bulky hydrophobic amino acids (Phe, Trp, Tyr, Leu, Ile) are recognized preferentially and thus considered destabilizing degrons since they allow faster degradation of their proteins.{{cite journal | vauthors = Tasaki T, Sriram SM, Park KS, Kwon YT | title = The N-end rule pathway | journal = Annual Review of Biochemistry | volume = 81 | pages = 261–89 | date = 2012 | pmid = 22524314 | pmc = 3610525 | doi = 10.1146/annurev-biochem-051710-093308 }}

File:Phosphodegron binding by ubiquitin ligase.png

A degron can be converted into its active form by a post-translational modification{{cite journal | vauthors = Herhaus L, Dikic I | title = Expanding the ubiquitin code through post-translational modification | journal = EMBO Reports | volume = 16 | issue = 9 | pages = 1071–83 | date = September 2015 | pmid = 26268526 | pmc = 4576978 | doi = 10.15252/embr.201540891 }} such as phosphorylation of a tyrosine, serine or threonine residue.{{cite journal | vauthors = Reinhardt HC, Yaffe MB | title = Phospho-Ser/Thr-binding domains: navigating the cell cycle and DNA damage response | journal = Nature Reviews Molecular Cell Biology | volume = 14 | issue = 9 | pages = 563–80 | date = September 2013 | pmid = 23969844 | doi = 10.1038/nrm3640 | s2cid = 149598 }} In this case, the ubiquitin ligase exclusively recognizes the phosphorylated version of the substrate due to stabilization within the binding site. For example, FBW7, the F-box substrate recognition unit of an SCF^FBW7ubiquitin ligase, stabilizes a phosphorylated substrate by hydrogen binding its arginine residues to the phosphate, as shown in the figure to the right. In absence of the phosphate, residues of FBW7 repel the substrate.

The presence of oxygen or other small molecules can influence degron recognition. The von Hippel-Lindau (VHL) protein (substrate recognition part of a specific E3 ligase), for instance, recognizes the hypoxia-inducible factor alpha (HIF-α) only under normal oxygen conditions, when its proline is hydroxylated. Under hypoxia, on the other hand, HIF-a is not hydroxylated, evades ubiquitination and thus operates in the cell at higher concentrations which can initiate transcriptional response to hypoxia.{{cite journal | vauthors = Jaakkola P, Mole DR, Tian YM, Wilson MI, Gielbert J, Gaskell SJ, von Kriegsheim A, Hebestreit HF, Mukherji M, Schofield CJ, Maxwell PH, Pugh CW, Ratcliffe PJ | title = Targeting of HIF-alpha to the von Hippel-Lindau ubiquitylation complex by O2-regulated prolyl hydroxylation | journal = Science | volume = 292 | issue = 5516 | pages = 468–72 | date = April 2001 | pmid = 11292861 | doi = 10.1126/science.1059796 | bibcode = 2001Sci...292..468J | s2cid = 20914281 | doi-access = free }} Another example of small molecule control of protein degradation is phytohormone auxin in plants.{{cite journal | vauthors = Shabek N, Zheng N | title = Plant ubiquitin ligases as signaling hubs | journal = Nature Structural & Molecular Biology | volume = 21 | issue = 4 | pages = 293–6 | date = April 2014 | pmid = 24699076 | doi = 10.1038/nsmb.2804 | s2cid = 41227590 }} Auxin binds to TIR1 (the substrate recognition domain of SCF^TIR1ubiquitin ligase) increasing the affinity of TIR1 for its substrates (transcriptional repressors: Aux/IAA), and promoting their degradation.

In addition to recognizing amino acids, ubiquitin ligases can also detect unusual features on substrates that serve as signals for their destruction. For example, San1 (Sir antagonist 1), a nuclear protein quality control in yeast, has a disordered substrate binding domain, which allows it to bind to hydrophobic domains of misfolded proteins. Misfolded or excess unassembled glycoproteins of the ERAD pathway, on the other hand, are recognized by Fbs1 and Fbs2, mammalian F-box proteins of E3 ligases SCF^Fbs1and SCF^Fbs2.{{cite journal | vauthors = Yoshida Y, Mizushima T, Tanaka K | title = Sugar-Recognizing Ubiquitin Ligases: Action Mechanisms and Physiology | journal = Frontiers in Physiology | volume = 10 | pages = 104 | date = 2019-02-19 | pmid = 30837888 | pmc = 6389600 | doi = 10.3389/fphys.2019.00104 | doi-access = free }} These recognition domains have small hydrophobic pockets allowing them to bind high-mannose containing glycans.

In addition to linear degrons, the E3 ligase can in some cases also recognize structural motifs on the substrate. In this case, the 3D motif can allow the substrate to directly relate its biochemical function to ubiquitination. This relation can be demonstrated with TRF1 protein (regulator of human telomere length), which is recognized by its corresponding E3 ligase (FBXO4) via an intermolecular beta sheet interaction. TRF1 cannot be ubiquinated while telomere bound, likely because the same TRF1 domain that binds to its E3 ligase also binds to telomeres.

E3 ubiquitin ligases regulate homeostasis, cell cycle, and DNA repair pathways, and as a result, a number of these proteins are involved in a variety of cancers, including famously MDM2, BRCA1, and Von Hippel-Lindau tumor suppressor.{{cite journal | vauthors = Lipkowitz S, Weissman AM | title = RINGs of good and evil: RING finger ubiquitin ligases at the crossroads of tumour suppression and oncogenesis | journal = Nature Reviews. Cancer | volume = 11 | issue = 9 | pages = 629–43 | date = August 2011 | pmid = 21863050 | pmc = 3542975 | doi = 10.1038/nrc3120 }} For example, a mutation of MDM2 has been found in stomach cancer,{{cite journal | vauthors = Hou YC, Deng JY | title = Role of E3 ubiquitin ligases in gastric cancer | journal = World Journal of Gastroenterology | volume = 21 | issue = 3 | pages = 786–93 | date = January 2015 | pmid = 25624711 | pmc = 4299330 | doi = 10.3748/wjg.v21.i3.786 | doi-access = free }} renal cell carcinoma,{{cite journal | vauthors = de Martino M, Taus C, Wessely IS, Lucca I, Hofbauer SL, Haitel A, Shariat SF, Klatte T | title = The T309G murine double minute 2 gene polymorphism is an independent prognostic factor for patients with renal cell carcinoma | journal = DNA and Cell Biology | volume = 34 | issue = 2 | pages = 107–12 | date = February 2015 | pmid = 25415135 | doi = 10.1089/dna.2014.2653 }} and liver cancer{{cite journal | vauthors = Tang T, Song X, Yang Z, Huang L, Wang W, Tan H | title = Association between murine double minute 2 T309G polymorphism and risk of liver cancer | journal = Tumour Biology | volume = 35 | issue = 11 | pages = 11353–7 | date = November 2014 | pmid = 25119589 | doi = 10.1007/s13277-014-2432-9 | s2cid = 16385927 }} (amongst others) to deregulate MDM2 concentrations by increasing its promoter’s affinity for the Sp1 transcription factor, causing increased transcription of MDM2 mRNA. Several proteomics-based experimental techniques are available for identifying E3 ubiquitin ligase-substrate pairs,{{cite journal | vauthors = Rayner SL, Morsch M, Molloy MP, Shi B, Chung R, Lee A | title = Using proteomics to identify ubiquitin ligase-substrate pairs: how novel methods may unveil therapeutic targets for neurodegenerative diseases | journal = Cellular and Molecular Life Sciences | volume = 76 | issue = 13 | pages = 2499–2510 | date = July 2019 | pmid = 30919022 | doi = 10.1007/s00018-019-03082-9 | s2cid = 85527795 | pmc = 11105231 }} such as proximity-dependent biotin identification (BioID), ubiquitin ligase-substrate trapping, and tandem ubiquitin-binding entities (TUBEs).

• A RING (Really Interesting New Gene) domain binds the E2 conjugase and might be found to mediate enzymatic activity in the E2-E3 complex{{cite journal | vauthors = Ardley HC, Robinson PA | title = E3 ubiquitin ligases | journal = Essays in Biochemistry | volume = 41 | pages = 15–30 | year = 2005 | pmid = 16250895 | doi = 10.1042/EB0410015 }}
• An F-box domain (as in the SCF complex) binds the ubiquitinated substrate. (e.g., Cdc 4, which binds the target protein Sic1; Grr1, which binds Cln).{{cite journal | vauthors = Bai C, Sen P, Hofmann K, Ma L, Goebl M, Harper JW, Elledge SJ | title = SKP1 connects cell cycle regulators to the ubiquitin proteolysis machinery through a novel motif, the F-box | journal = Cell | volume = 86 | issue = 2 | pages = 263–74 | date = July 1996 | pmid = 8706131 | doi = 10.1016/S0092-8674(00)80098-7 | doi-access = free }}
• A HECT domain, which is involved in the transfer of ubiquitin from the E2 to the substrate.

In 2001, work from the labs of Craig Crews and Raymond Deshaies described the development of proteolysis-targeting chimeras (PROTACs).{{Cite journal |last1=Sakamoto |first1=K. M. |last2=Kim |first2=K. B. |last3=Kumagai |first3=A. |last4=Mercurio |first4=F. |last5=Crews |first5=C. M. |last6=Deshaies |first6=R. J. |date=2001-07-17 |title=Protacs: chimeric molecules that target proteins to the Skp1-Cullin-F box complex for ubiquitination and degradation |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=98 |issue=15 |pages=8554–8559 |doi=10.1073/pnas.141230798 |doi-access=free |issn=0027-8424 |pmc=37474 |pmid=11438690|bibcode=2001PNAS...98.8554S }} Using a small molecule to recruit an E3 ubiquitin ligase to a target protein, this work demonstrated that induced proximity could be used to effect the ubiquitination and proteasomal degradation of a target protein. PROTACs have been frequently applied using the E3 ubiquitin ligases CRBN{{Cite journal |last1=Lu |first1=Jing |last2=Qian |first2=Yimin |last3=Altieri |first3=Martha |last4=Dong |first4=Hanqing |last5=Wang |first5=Jing |last6=Raina |first6=Kanak |last7=Hines |first7=John |last8=Winkler |first8=James D. |last9=Crew |first9=Andrew P. |last10=Coleman |first10=Kevin |last11=Crews |first11=Craig M. |date=2015-06-18 |title=Hijacking the E3 Ubiquitin Ligase Cereblon to Efficiently Target BRD4 |journal=Chemistry & Biology |volume=22 |issue=6 |pages=755–763 |doi=10.1016/j.chembiol.2015.05.009 |issn=1879-1301 |pmc=4475452 |pmid=26051217}}{{Cite journal |last1=Winter |first1=Georg E. |last2=Buckley |first2=Dennis L. |last3=Paulk |first3=Joshiawa |last4=Roberts |first4=Justin M. |last5=Souza |first5=Amanda |last6=Dhe-Paganon |first6=Sirano |last7=Bradner |first7=James E. |date=2015-06-19 |title=Phthalimide conjugation as a strategy for in vivo target protein degradation |journal=Science |language=en |volume=348 |issue=6241 |pages=1376–1381 |doi=10.1126/science.aab1433 |issn=0036-8075 |pmc=4937790 |pmid=25999370|bibcode=2015Sci...348.1376W }} and VHL{{Cite journal |last1=Bondeson |first1=Daniel P. |last2=Mares |first2=Alina |last3=Smith |first3=Ian E. D. |last4=Ko |first4=Eunhwa |last5=Campos |first5=Sebastien |last6=Miah |first6=Afjal H. |last7=Mulholland |first7=Katie E. |last8=Routly |first8=Natasha |last9=Buckley |first9=Dennis L. |last10=Gustafson |first10=Jeffrey L. |last11=Zinn |first11=Nico |last12=Grandi |first12=Paola |last13=Shimamura |first13=Satoko |last14=Bergamini |first14=Giovanna |last15=Faelth-Savitski |first15=Maria |date=August 2015 |title=Catalytic in vivo protein knockdown by small-molecule PROTACs |journal=Nature Chemical Biology |volume=11 |issue=8 |pages=611–617 |doi=10.1038/nchembio.1858 |issn=1552-4469 |pmc=4629852 |pmid=26075522}}{{Cite journal |last1=Zengerle |first1=Michael |last2=Chan |first2=Kwok-Ho |last3=Ciulli |first3=Alessio |date=2015-08-21 |title=Selective Small Molecule Induced Degradation of the BET Bromodomain Protein BRD4 |url=https://pubs.acs.org/doi/10.1021/acschembio.5b00216?ref=recommended& |journal=ACS Chemical Biology |volume=10 |issue=8 |pages=1770–1777 |doi=10.1021/acschembio.5b00216 |issn=1554-8929|pmc=4548256 }} to degrade various targets of biological and therapeutic relevance. Multiple groups have sought out additional E3 ligases to co-opt for targeted protein degradation such as FBXO22{{Cite journal |last1=Nie |first1=David Y. |last2=Tabor |first2=John R. |last3=Li |first3=Jianping |last4=Kutera |first4=Maria |last5=St-Germain |first5=Jonathan |last6=Hanley |first6=Ronan P. |last7=Wolf |first7=Esther |last8=Paulakonis |first8=Ethan |last9=Kenney |first9=Tristan M. G. |last10=Duan |first10=Shili |last11=Shrestha |first11=Suman |last12=Owens |first12=Dominic D. G. |last13=Maitland |first13=Matthew E. R. |last14=Pon |first14=Ailing |last15=Szewczyk |first15=Magdalena |date=December 2024 |title=Recruitment of FBXO22 for targeted degradation of NSD2 |journal=Nature Chemical Biology |volume=20 |issue=12 |pages=1597–1607 |doi=10.1038/s41589-024-01660-y |issn=1552-4469 |pmc=11581931 |pmid=38965384|pmc-embargo-date=December 1, 2025 }}{{Cite journal |last1=Basu |first1=Ananya A. |last2=Zhang |first2=Chenlu |last3=Riha |first3=Isabella A. |last4=Magassa |first4=Assa |last5=Campos |first5=Miguel A. |last6=Caldwell |first6=Alana G. |last7=Ko |first7=Felicia |last8=Zhang |first8=Xiaoyu |date=December 2024 |title=A CRISPR activation screen identifies FBXO22 supporting targeted protein degradation |journal=Nature Chemical Biology |volume=20 |issue=12 |pages=1608–1616 |doi=10.1038/s41589-024-01655-9 |issn=1552-4469 |pmc=11581908 |pmid=38965383|pmc-embargo-date=June 1, 2025 }}{{Cite journal |last1=Kagiou |first1=Chrysanthi |last2=Cisneros |first2=Jose A. |last3=Farnung |first3=Jakob |last4=Liwocha |first4=Joanna |last5=Offensperger |first5=Fabian |last6=Dong |first6=Kevin |last7=Yang |first7=Ka |last8=Tin |first8=Gary |last9=Horstmann |first9=Christina S. |last10=Hinterndorfer |first10=Matthias |last11=Paulo |first11=Joao A. |last12=Scholes |first12=Natalie S. |last13=Sanchez Avila |first13=Juan |last14=Fellner |first14=Michaela |last15=Andersch |first15=Florian |date=2024-06-26 |title=Alkylamine-tethered molecules recruit FBXO22 for targeted protein degradation |journal=Nature Communications |language=en |volume=15 |issue=1 |pages=5409 |doi=10.1038/s41467-024-49739-3 |pmid=38926334 |pmc=11208438 |bibcode=2024NatCo..15.5409K |issn=2041-1723}} and KLHDC2.{{Cite journal |last1=Scott |first1=Daniel C. |last2=Dharuman |first2=Suresh |last3=Griffith |first3=Elizabeth |last4=Chai |first4=Sergio C. |last5=Ronnebaum |first5=Jarrid |last6=King |first6=Moeko T. |last7=Tangallapally |first7=Rajendra |last8=Lee |first8=Chan |last9=Gee |first9=Clifford T. |last10=Yang |first10=Lei |last11=Li |first11=Yong |last12=Loudon |first12=Victoria C. |last13=Lee |first13=Ha Won |last14=Ochoada |first14=Jason |last15=Miller |first15=Darcie J. |date=2024-10-12 |title=Principles of paralog-specific targeted protein degradation engaging the C-degron E3 KLHDC2 |url=https://www.nature.com/articles/s41467-024-52966-3 |journal=Nature Communications |language=en |volume=15 |issue=1 |pages=8829 |doi=10.1038/s41467-024-52966-3 |bibcode=2024NatCo..15.8829S |issn=2041-1723|pmc=11470957 }}

While PROTACs generally are heterobifunctional compounds linking an E3 ligase binder to a target protein binder, molecular glues also exist that induce protein-protein interactions with E3 ligases, leading to degradation of various substrate proteins. Molecular glues often have been discovered through serendipity,{{Cite journal |last1=Ito |first1=Takumi |last2=Ando |first2=Hideki |last3=Suzuki |first3=Takayuki |last4=Ogura |first4=Toshihiko |last5=Hotta |first5=Kentaro |last6=Imamura |first6=Yoshimasa |last7=Yamaguchi |first7=Yuki |last8=Handa |first8=Hiroshi |date=2010-03-12 |title=Identification of a Primary Target of Thalidomide Teratogenicity |url=https://www.science.org/doi/10.1126/science.1177319 |journal=Science |volume=327 |issue=5971 |pages=1345–1350 |doi=10.1126/science.1177319|pmid=20223979 |bibcode=2010Sci...327.1345I }}{{Cite journal |last1=Choi |first1=J. |last2=Chen |first2=J. |last3=Schreiber |first3=S. L. |last4=Clardy |first4=J. |date=1996-07-12 |title=Structure of the FKBP12-rapamycin complex interacting with the binding domain of human FRAP |url=https://pubmed.ncbi.nlm.nih.gov/8662507 |journal=Science (New York, N.Y.) |volume=273 |issue=5272 |pages=239–242 |doi=10.1126/science.273.5272.239 |issn=0036-8075 |pmid=8662507|bibcode=1996Sci...273..239C }}{{Cite journal |last1=Liu |first1=Jun |last2=Farmer |first2=Jesse D. |last3=Lane |first3=Willam S. |last4=Friedman |first4=Jeff |last5=Weissman |first5=Irving |last6=Schreiber |first6=Stuart L. |date=1991-08-23 |title=Calcineurin is a common target of cyclophilin-cyclosporin A and FKBP-FK506 complexes |url=https://www.cell.com/cell/abstract/0092-8674(91)90124-H |journal=Cell |language=English |volume=66 |issue=4 |pages=807–815 |doi=10.1016/0092-8674(91)90124-H |issn=0092-8674 |pmid=1715244}} though various methodologies have been explored to expedite the discovery of molecular glues.{{Citation |last1=Bushman |first1=Jonathan W. |title=Discovery of a VHL molecular glue degrader of GEMIN3 by Picowell RNA-seq |date=2025-03-19 |url=https://www.biorxiv.org/content/10.1101/2025.03.19.644003v1 |access-date=2025-04-07 |publisher=bioRxiv |language=en |doi=10.1101/2025.03.19.644003 |last2=Deng |first2=Weixian |last3=Samarasinghe |first3=Kusal T. G. |last4=Liu |first4=Han-Yuan |last5=Li |first5=Shiqian |last6=Vaish |first6=Amit |last7=Golkar |first7=Alex |last8=Ou |first8=Shu-Ching |last9=Ahn |first9=Jinwoo}}{{Cite journal |last1=Razumkov |first1=Hlib |last2=Jiang |first2=Zixuan |last3=Baek |first3=Kheewoong |last4=You |first4=Inchul |last5=Geng |first5=Qixiang |last6=Donovan |first6=Katherine A. |last7=Tang |first7=Michelle T. |last8=Metivier |first8=Rebecca J. |last9=Mageed |first9=Nada |last10=Seo |first10=Pooreum |last11=Li |first11=Zhengnian |last12=Byun |first12=Woong Sub |last13=Hinshaw |first13=Stephen M. |last14=Sarott |first14=Roman C. |last15=Fischer |first15=Eric S. |date=2024-11-20 |title=Discovery of CRBN-Dependent WEE1 Molecular Glue Degraders from a Multicomponent Combinatorial Library |journal=Journal of the American Chemical Society |volume=146 |issue=46 |pages=31433–31443 |doi=10.1021/jacs.4c06127 |issn=0002-7863 |pmc=11800961 |pmid=39499896|pmc-embargo-date=May 20, 2025 |bibcode=2024JAChS.14631433R }}{{Citation |last1=Zhuang |first1=Zhe |title=Charged Molecular Glue Discovery Enabled by Targeted Degron Display |date=2024-09-26 |url=https://www.biorxiv.org/content/10.1101/2024.09.24.614843v1 |access-date=2025-04-07 |publisher=bioRxiv |language=en |doi=10.1101/2024.09.24.614843 |last2=Byun |first2=Woong Sub |last3=Chrustowicz |first3=Jakub |last4=Kozicka |first4=Zuzanna |last5=Li |first5=Veronica L. |last6=Abeja |first6=Dinah M. |last7=Donovan |first7=Katherine A. |last8=Sepic |first8=Sara |last9=You |first9=Inchul}}{{Cite journal |last1=Wang |first1=Zefeng |last2=Shaabani |first2=Shabnam |last3=Gao |first3=Xiang |last4=Ng |first4=Yuen Lam Dora |last5=Sapozhnikova |first5=Valeriia |last6=Mertins |first6=Philipp |last7=Krönke |first7=Jan |last8=Dömling |first8=Alexander |date=2023-12-19 |title=Direct-to-biology, automated, nano-scale synthesis, and phenotypic screening-enabled E3 ligase modulator discovery |url=https://www.nature.com/articles/s41467-023-43614-3 |journal=Nature Communications |language=en |volume=14 |issue=1 |pages=8437 |doi=10.1038/s41467-023-43614-3 |pmid=38114468 |bibcode=2023NatCo..14.8437W |issn=2041-1723}}{{Cite journal |last1=Toriki |first1=Ethan S. |last2=Papatzimas |first2=James W. |last3=Nishikawa |first3=Kaila |last4=Dovala |first4=Dustin |last5=Frank |first5=Andreas O. |last6=Hesse |first6=Matthew J. |last7=Dankova |first7=Daniela |last8=Song |first8=Jae-Geun |last9=Bruce-Smythe |first9=Megan |last10=Struble |first10=Heidi |last11=Garcia |first11=Francisco J. |last12=Brittain |first12=Scott M. |last13=Kile |first13=Andrew C. |last14=McGregor |first14=Lynn M. |last15=McKenna |first15=Jeffrey M. |date=2023-05-24 |title=Rational Chemical Design of Molecular Glue Degraders |journal=ACS Central Science |volume=9 |issue=5 |pages=915–926 |doi=10.1021/acscentsci.2c01317 |issn=2374-7943 |pmc=10214506 |pmid=37252349}}{{Cite journal |last1=Baek |first1=Kheewoong |last2=Metivier |first2=Rebecca J. |last3=Roy Burman |first3=Shourya S. |last4=Bushman |first4=Jonathan W. |last5=Yoon |first5=Hojong |last6=Lumpkin |first6=Ryan J. |last7=Abeja |first7=Dinah M. |last8=Lakshminarayan |first8=Megha |last9=Yue |first9=Hong |last10=Ojeda |first10=Samuel |last11=Verano |first11=Alyssa L. |last12=Gray |first12=Nathanael S. |last13=Donovan |first13=Katherine A. |last14=Fischer |first14=Eric S. |date=2024-10-04 |title=Unveiling the hidden interactome of CRBN molecular glues with chemoproteomics |journal=BioRxiv: The Preprint Server for Biology |pages=2024.09.11.612438 |doi=10.1101/2024.09.11.612438 |issn=2692-8205 |pmc=11419069 |pmid=39314457}}

Biologic modalities for targeted protein degradation have also been explored by fusing E3 ligases to target recognition domains such as nanobodies. These modalities are sometimes referred to as bioPROTACs.{{Cite journal |last1=Lim |first1=Shuhui |last2=Khoo |first2=Regina |last3=Peh |first3=Khong Ming |last4=Teo |first4=Jinkai |last5=Chang |first5=Shih Chieh |last6=Ng |first6=Simon |last7=Beilhartz |first7=Greg L. |last8=Melnyk |first8=Roman A. |last9=Johannes |first9=Charles W. |last10=Brown |first10=Christopher J. |last11=Lane |first11=David P. |last12=Henry |first12=Brian |last13=Partridge |first13=Anthony W. |date=2020-03-17 |title=bioPROTACs as versatile modulators of intracellular therapeutic targets including proliferating cell nuclear antigen (PCNA) |journal=Proceedings of the National Academy of Sciences |volume=117 |issue=11 |pages=5791–5800 |doi=10.1073/pnas.1920251117 |doi-access=free |pmc=7084165 |pmid=32123106|bibcode=2020PNAS..117.5791L }}{{Cite journal |last1=Fletcher |first1=Alice |last2=Clift |first2=Dean |last3=de Vries |first3=Emma |last4=Martinez Cuesta |first4=Sergio |last5=Malcolm |first5=Timothy |last6=Meghini |first6=Francesco |last7=Chaerkady |first7=Raghothama |last8=Wang |first8=Junmin |last9=Chiang |first9=Abby |last10=Weng |first10=Shao Huan Samuel |last11=Tart |first11=Jonathan |last12=Wong |first12=Edmond |last13=Donohoe |first13=Gerard |last14=Rawlins |first14=Philip |last15=Gordon |first15=Euan |date=2023-11-04 |title=A TRIM21-based bioPROTAC highlights the therapeutic benefit of HuR degradation |journal=Nature Communications |language=en |volume=14 |issue=1 |pages=7093 |doi=10.1038/s41467-023-42546-2 |pmid=37925433 |pmc=10625600 |bibcode=2023NatCo..14.7093F |issn=2041-1723}} While bioPROTACs are advantageous for targeting proteins lacking small molecule ligands, challenges in delivery, pharmacokinetics, and immunogenicity have so far precluded clinical development.{{Cite journal |last1=Ou |first1=Lisha |last2=Setegne |first2=Mekedlawit T. |last3=Elliot |first3=Jeandele |last4=Shen |first4=Fangfang |last5=Dassama |first5=Laura M. K. |date=2025-02-26 |title=Protein-Based Degraders: From Chemical Biology Tools to Neo-Therapeutics |journal=Chemical Reviews |volume=125 |issue=4 |pages=2120–2183 |doi=10.1021/acs.chemrev.4c00595 |issn=1520-6890 |pmc=11870016 |pmid=39818743|pmc-embargo-date=January 17, 2026 }} Studies exploring different delivery mechanisms have sought to address these shortcomings.{{Cite journal |last1=Chan |first1=Alexander |last2=Haley |first2=Rebecca M. |last3=Najar |first3=Mohd Altaf |last4=Gonzalez-Martinez |first4=David |last5=Bugaj |first5=Lukasz J. |last6=Burslem |first6=George M. |last7=Mitchell |first7=Michael J. |last8=Tsourkas |first8=Andrew |date=2024-07-10 |title=Lipid-mediated intracellular delivery of recombinant bioPROTACs for the rapid degradation of undruggable proteins |url=https://www.nature.com/articles/s41467-024-50235-x |journal=Nature Communications |language=en |volume=15 |issue=1 |pages=5808 |doi=10.1038/s41467-024-50235-x |pmid=38987546 |bibcode=2024NatCo..15.5808C |issn=2041-1723|pmc=11237011 }} In another variant of this idea, bispecific antibodies to recruit membrane-bound E3 ligases to cell surface proteins (AbTACs) have also been developed.{{Cite journal |last1=Cotton |first1=Adam D. |last2=Nguyen |first2=Duy P. |last3=Gramespacher |first3=Josef A. |last4=Seiple |first4=Ian B. |last5=Wells |first5=James A. |date=2021-01-20 |title=Development of Antibody-Based PROTACs for the Degradation of the Cell-Surface Immune Checkpoint Protein PD-L1 |journal=Journal of the American Chemical Society |volume=143 |issue=2 |pages=593–598 |doi=10.1021/jacs.0c10008 |issn=0002-7863 |pmc=8154509 |pmid=33395526|bibcode=2021JAChS.143..593C }}

{{div col|colwidth=20em}}

• E3A
• mdm2
• Anaphase-promoting complex (APC)
• UBR5 (EDD1)
• SOCS/ BC-box/ eloBC/ CUL5/ RING
• LNXp80
• CBX4, CBLL1
• HACE1
• HECTD1, HECTD2, HECTD3, HECTD4
• HECW1, HECW2
• HERC1, HERC2, HERC3, HERC4, HERC5, HERC6
• HUWE1, ITCH
• NEDD4, NEDD4L
• PPIL2
• PRPF19
• PIAS1, PIAS2, PIAS3, PIAS4
• RANBP2
• RNF4, RNF167
• RBX1
• SMURF1, SMURF2
• STUB1
• TOPORS
• TRIP12
• UBE3A, UBE3B, UBE3C, UBE3D
• UBE4A, UBE4B
• UBOX5
• UBR5
• VHL
• WWP1, WWP2
• Parkin
• MKRN1

{{Div col end}}

• ERAD
• Ubiquitin
• Ubiquitin-activating enzyme
• Ubiquitin-conjugating enzyme

{{Reflist|33em}}

• [http://www.ebi.ac.uk/pdbe-apps/quips?story=LordCBL Quips article describing E3 Ligase function] {{Webarchive|url=https://web.archive.org/web/20121130051453/http://www.ebi.ac.uk/pdbe-apps/quips?story=LordCBL |date=2012-11-30 }} at [http://www.pdbe.org PDBe]
• {{MeshName|Ubiquitin-Protein+Ligases}}
• {{EC number|6.3.2.19}}

{{Posttranslational modification}}

{{Ligases CO CS and CN}}

{{Enzymes}}

{{Portal bar|Biology|border=no}}

{{DEFAULTSORT:Ubiquitin Ligase}}

Category:EC 6.3

Category:Post-translational modification