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Information on EC 2.3.2.26 - HECT-type E3 ubiquitin transferase and Organism(s) Homo sapiens and UniProt Accession P46934

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EC Tree
     2 Transferases
         2.3 Acyltransferases
             2.3.2 Aminoacyltransferases
                2.3.2.26 HECT-type E3 ubiquitin transferase
IUBMB Comments
In the first step the enzyme transfers ubiquitin from the E2 ubiquitin-conjugating enzyme (EC 2.3.2.23) to a cysteine residue in its HECT domain (which is located in the C-terminal region), forming a thioester bond. In a subsequent step the enzyme transfers the ubiquitin to an acceptor protein, resulting in the formation of an isopeptide bond between the C-terminal glycine residue of ubiquitin and the epsilon-amino group of an L-lysine residue of the acceptor protein. cf. EC 2.3.2.27, RING-type E3 ubiquitin transferase and EC 2.3.2.31, RBR-type E3 ubiquitin transferase.
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Homo sapiens
UNIPROT: P46934
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Word Map
The taxonomic range for the selected organisms is: Homo sapiens
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
Reaction Schemes
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[E2 ubiquitin-conjugating enzyme]-S-ubiquitinyl-L-cysteine
+
[acceptor protein]-L-lysine
=
[E2 ubiquitin-conjugating enzyme]-L-cysteine
+
[acceptor protein]-N6-ubiquitinyl-L-lysine
Synonyms
nedd4, smurf1, smurf2, huwe1, nedd4l, nedd4-1, e6-ap, ube3c, trp120, trip12, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E3 ligase
-
E3 ubiquitin-protein ligase NEDD4
-
HECT ligase
-
Nedd4 HECT
-
apoptosis-resistant E3 ligase 1
-
atrophin-1 interacting protein 2
-
HECT domain E3 ubiquitin ligase
-
HECT-domain ubiquitin ligase
-
Nedd4L
ubiquitin-protein ligase E3C
-
WW domain-containing E3 Ub–protein ligase 2
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
[E2 ubiquitin-conjugating enzyme]-S-ubiquitinyl-L-cysteine + [acceptor protein]-L-lysine = [E2 ubiquitin-conjugating enzyme]-L-cysteine + [acceptor protein]-N6-ubiquitinyl-L-lysine
show the reaction diagram
HECT ligases directly catalyse protein ubiquitination and non-covalently interact with ubiquitin. The ubiquitin bindung surface on the HECT might act to bind a ubiquitin moiety that is already conjugated to a protein substrate, thus promoting polyubiquitination. Mutation in the ubiquitin bindung surface strongly impairs free-chain fdormation and ubiquitination of all substrates tested
PATHWAY SOURCE
PATHWAYS
-
-
SYSTEMATIC NAME
IUBMB Comments
[E2 ubiquitin-conjugating enzyme]-S-ubiquitinyl-L-cysteine:[acceptor protein] ubiquitin transferase (isopeptide bond-forming)
In the first step the enzyme transfers ubiquitin from the E2 ubiquitin-conjugating enzyme (EC 2.3.2.23) to a cysteine residue in its HECT domain (which is located in the C-terminal region), forming a thioester bond. In a subsequent step the enzyme transfers the ubiquitin to an acceptor protein, resulting in the formation of an isopeptide bond between the C-terminal glycine residue of ubiquitin and the epsilon-amino group of an L-lysine residue of the acceptor protein. cf. EC 2.3.2.27, RING-type E3 ubiquitin transferase and EC 2.3.2.31, RBR-type E3 ubiquitin transferase.
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
S-ubiquitinyl-[E2 ubiquitin-conjugating enzyme]-L-cysteine + [Spry2]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + N6-ubiquitinyl-[Spry2]-L-lysine
show the reaction diagram
Spry2 is a regulator of receptor tyrosine kinase signaling in development and disease
isoform Nedd4 polyubiquitinates Spry2 via Lys48 on ubiquitin and decreases its stability. The Spry2/Nedd4 association involves theWW domains of Nedd4 and requires phosphorylation of the Mnk2 kinase sites, Ser112 and Ser121, on Spry2. The phospho-Ser112/121 region on Spry2 that binds WW domains of Nedd4 is a non-canonical WW domain binding region that does not contain Pro residues after phospho-Ser
-
?
[HECT-E3-ubiquitin-carrier protein NEDD4]-S-ubiquitin-L-cysteine + [gamma-epithel Na+-channel]-L-lysine
[HECT-E3-ubiquitin-carrier protein NEDD4]-L-cysteine + [gamma-epithel Na+-channel]-N6-ubiquinyl-L-lysine
show the reaction diagram
His-tagged Ube2D3, in addition the reaction mixture contains purified E1 enzyme and ubiquitin
-
-
?
[HECT-E3-ubiquitin-carrier protein NEDD4]-S-ubiquitin-L-cysteine + [SQSTM1]-L-lysine
[HECT-E3-ubiquitin-carrier protein NEDD4]-L-cysteine + [SQSTM1]-N6-ubiquinyl-L-lysine
show the reaction diagram
SQSTM1 i.e. an autophagic cargo receptor involved in selective autophagy
-
-
?
[Nedd4-1-ubiquitin-conjugating enzyme E2]-S-ubiquitin-L-cysteine + [activated Cdc42-associated tyrosine kinase]-L-lysine
[Nedd4-1ubiquitin-conjugating enzyme E2]-L-cysteine + [activated Cdc42-associated tyrosine kinase]-N6-ubiquitinyl-L-lysine
show the reaction diagram
activated Cdc42-associated tyrosine kinase is ubiquitinated by HECT E3 ubiquitin ligase Nedd4-1 and degraded along with epidermal growth factor receptor in response to epidermal growth factor stimulation. Activated Cdc42-associated tyrosine kinase interacts with Nedd4-1 through a conserved PPXY WW-binding motif. The WW3 domain in Nedd4-1 is critical for binding to activated Cdc42-associated tyrosine kinase. Deletion of the sterile alpha motif SAM-domain at the N-terminus dramatically reduces the ubiquitination of activated Cdc42-associated tyrosine kinase by Nedd4-1, while deletion of the Uba domain dramatically enhances the ubiquitination. Activated Cdc42-associated tyrosine kinase degradation is processed by lysosomes, not proteasomes
-
-
?
[Nedd4-1-ubiquitin-conjugating enzyme E2]-S-ubiquitin-L-cysteine + [epidermal growth factor receptor]-L-lysine
[Nedd4-1-ubiquitin-conjugating enzyme E2]-L-cysteine + [epidermal growth factor receptor]-N6-ubiquitinyl-L-lysine
show the reaction diagram
epidermal growth factor receptor and activated Cdc42-associated tyrosine kinase are ubiquitinated by ubiquitin ligase Nedd4-1 in response to epidermal growth factor stimulation
-
-
?
[ubiquitin-conjugating enzyme E2D3]-S-ubiquitin-L-cysteine + [latent membrane protein 2A LMP2A]-L-lysine
[ubiquitin-conjugating enzyme E2D3]-L-cysteine + [latent membrane protein 2A LMP2A]-N6-ubiquitinyl-L-lysine
show the reaction diagram
His-tagged Ube2D3, in addition the reaction mixture contains purified E1 enzyme and ubiquitin
-
-
?
S-(ubiquitin)n-[E2 ubiquitin-conjugating enzyme]-L-cysteine + [adenomatous polyposis coli]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + N6-(ubiquitin)n-[adenomatous polyposis coli]-L-lysine
show the reaction diagram
-
adenomatous polyposis coli protein functions as a negative regulator of the Wnt signaling pathway
isoform HECTD1 modifies adenomatous polyposis coli with Lys63 polyubiquitin
-
?
S-(ubiquitin)n-[E2 ubiquitin-conjugating enzyme]-L-cysteine + [Dvl2]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + N6-(ubiquitin)n-[Dvl2]-L-lysine
show the reaction diagram
Dvl2 i.e. dishevelled, a central mediator for both Wnt/beta-catenin and Wnt/planar cell polarity pathways
isoform NEDD4L mediates polyubiquitination of Dvl2 at Lys6, Lys27, and Lys29
-
?
S-ubiquitinyl-[E2 ubiquitin-conjugating enzyme]-L-cysteine + [caspase-8]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + N6-ubiquitinyl-[caspase-8]-L-lysine
show the reaction diagram
-
isoform HECTD3 ubiquitinates caspase-8 with K63-linked polyubiquitin chains that do not target caspase-8 for degradation but decrease the caspase-8 activation
-
?
S-ubiquitinyl-[E2 ubiquitin-conjugating enzyme]-L-cysteine + [ING2]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + N6-ubiquitinyl-[ING2]-L-lysine
show the reaction diagram
ING2 i.e. candidate tumor suppressor Inhibitor of Growth 2
isoform Smurf1 interacts with and targets ING2 for poly-ubiquitination and proteasomal degradation. The ING2 binding domain in Smurf1 was mapped to the catalytic HECT domain. The C-terminal PHD domain of ING2 is required for Smurf1-mediated degradation
-
?
S-ubiquitinyl-[HECT-type E3 ubiquitin transferase]-L-cysteine + [Sox6 protein]-L-lysine
[HECT-type E3 ubiquitin transferase]-L-cysteine + N6-ubiquitinyl-[Sox6 protein]-L-lysine
show the reaction diagram
-
-
-
?
S-ubiquitinyl-[UbcH5a]-L-cysteine + [ubiquitin mutant G76V]-L-lysine
[UbcH5a]-L-cysteine + N6-ubiquitinyl-[mutant G76V]-L-lysine
show the reaction diagram
-
-
-
?
S-ubiquitinyl-[UbcH5a]-L-cysteine + [ubiquitin-DELTAGG]-L-lysine
[UbcH5a]-L-cysteine + N6-ubiquitinyl-[ubiquitin-DELTAGG]-L-lysine
show the reaction diagram
ubiquitin-DELTAGG i.e. mutant ubiquitin lacking the two C-terminal glycine residues, cannot be conjugated to other proteins
-
-
?
S-ubiquitinyl-[UbcH7]-L-cysteine + [endophilin A]-L-lysine
[UbcH7]-L-cysteine + N6-ubiquitinyl-[endophilin A]-L-lysine
show the reaction diagram
-
isoform Itch ubiquitinates SH3 domain-containing protein endophilin A1 and the SH3/proline-rich domain interaction facilitates this activity. EGF treatment of cells stimulates endophilin A1 ubiquitination
-
?
[AIP2-ubiquitin-conjugating enzyme E2]-S-ubiquitin-L-cysteine + [EGR2]-L-lysine
[AIP2-ubiquitin-conjugating enzyme E2]-L-cysteine + [EGR2]-N6-ubiquitinyl-L-lysine
show the reaction diagram
EGR2, a zinc finger transcription factor that has been found to regulate Fas ligand expression during activation-induced T-cell death
-
-
?
[E2 ubiquitin-conjugating enzyme]-S-ubiquitinyl-L-cysteine + [Dvl2]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + [Dvl2]-N6-ubiquitinyl-L-lysine
show the reaction diagram
-
-
-
?
[E2 ubiquitin-conjugating enzyme]-S-ubiquitinyl-L-cysteine + [Ubl4A]-L-lysine
[E2 ubiquitin-conjugating enzyme]-L-cysteine + [Ubl4A]-N6-ubiquitinyl-L-lysine
show the reaction diagram
Ubl4A, i.e. subunit of the Bag6 chaperone holdase complex. HUWE1 degrades unassembled Ubl4A in the cytoplasm
-
-
?
[HECT-E3-ubiquitin-carrier protein Arel1]-S-ubiquitin-L-cysteine + [SMAC]-L-lysine
[HECT-E3-ubiquitin-carrier protein Arel1]-L-cysteine + [SMAC]-N6-ubiquinyl-L-lysine
show the reaction diagram
SMAC i.e. proapoptotic protein second mitochondria-derived activator of caspase
-
-
?
[HECT-E3-ubiquitin-carrier protein NEDD4L]-S-ubiquitin-L-cysteine + [Ubc5B]-L-lysine
[HECT-E3-ubiquitin-carrier protein NEDD4]-L-cysteine + [Ubc5B]-N6-ubiquinyl-L-lysine
show the reaction diagram
-
-
-
-
?
[TRIP1-ubiquitin-conjugating enzyme E2]-S-ubiquitin-L-cysteine + [APP-BP1]-L-lysine
[TRIP12-ubiquitin-conjugating enzyme E2]-L-cysteine + [APP-BP1]-N6-ubiquitinyl-L-lysine
show the reaction diagram
ubiquitin ligase TRIP12 functions as an E3 enzyme of APP-BP1 and additionally requires an E4 activity for polyubiquitination of APP-BP1. APP-BP1 is part of the ubiquitin-like protein NEDD8 activating enzyme. TRIP12 specifically interacts with the APP-BP1 monomer but not with the APP-BP1/Uba3 heterodimer. Overexpression of TRIP12 enhances the degradation of APP-BP1, whereas knockdown of TRIP12 stabilizes it
-
-
?
[ubiquitin ligase HECTD3]-S-ubiquitin-L-cysteine + [Tara]-L-lysine
[ubiquitin ligase HECTD3]-L-cysteine + [Tara]-N6-ubiquitinyl-L-lysine
show the reaction diagram
Tara, Trio-associated repeat on actin, is an interacting partner of guanine nucleotide exchange factors Trio and TRF1. Ubiquitin-protein ligase HECTD3 directly binds Tara in vitro and forms a complex with Tara in vivo. Overexpression of HECTD3 enhances the ubiquitination of Tara in vivo and promotes the turnover of Tara, whereas depletion of HECTD3 by small interfering RNA decreases Tara degradation
-
-
?
additional information
?
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
S-ubiquitinyl-[HECT-type E3 ubiquitin transferase]-L-cysteine + [Sox6 protein]-L-lysine
[HECT-type E3 ubiquitin transferase]-L-cysteine + N6-ubiquitinyl-[Sox6 protein]-L-lysine
show the reaction diagram
-
-
-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4-[[(3-chloro-1,4-dioxo-1,4-dihydronaphthalen-2-yl)amino]methyl]-N-(pyridin-4-yl)benzamide
compound exhibits remarkable anticancer activity with tumor growth inhibition values of 98.3% and 100% at 25 mg/kg and 50 mg/kg orally daily, respectively, against human RPMI-8226 multiple myeloma xenograft. Treatment with the compound also shows a decrease of Itch level in human RPMI-8226 multiple myeloma cells
-
Calmodulin
UBE3B interacts with calmodulin via its N-terminal isoleucine-glutamine motif. Deletion of the motif (amino acids 29-58) results in loss of calmodulin binding and a significant increase in the in vitro ubiquitylation activity of UBE3B. Changes in calcium levels in vitro disrupt the calmodulin-UBE3B interaction
chlorpromazine
minimum inhibitory concentration is 0.3 mM
chlorprothixene
minimum inhibitory concentration is 0.3 mM
clomipramine
antidepressant drug, specifically blocks isoform ITCH auto-ubiquitylation, as well as p73 ubiquitylation. Treating a panel of breast, prostate and bladder cancer cell lines with clomipramine, or its homologs, leads to reduced cancer cell growth, and synergize with gemcitabine or mitomycin in killing cancer cells by blocking autophagy. Minimum inhibitory concentration is 0.3 mM
N-acetylphenylalanylamide
-
i.e. Phe-823 mimic, acts as a noncompetitive inhibitor of polyubiquitin chain elongation by destabilizing the active trimer
norclomipramine
minimum inhibitory concentration is 0.3 mM
additional information
autoinhibition mechanism of domains C2-HECT is not observed in isoform Smurf1
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ARRDC1
alpha-arrestin, anchoring an ARRDC1-GFP construct to the membrane results in efficient activation and autoubiquitination of NEDD4 ligase
-
Ca2+
when the concentration of calcium is increased, calmodulin is released from UBE3B, resulting in an increase in ubiquitylation activity
Ndfip2
adaptor protein containing multiple PY domains
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0046 - 0.037
[Ubc5B]-L-lysine
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000009 - 0.012
[Ubc5B]-L-lysine
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
8
N-acetylphenylalanylamide
-
wild-type, pH 7.5, 37°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.6
assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
assay at
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
isoform Itch co-localizes with markers of the endosomal system in a C2 domain-dependent manner and upon EGF stimulation, substrate endophilin A1 translocates to an EGF-positive endosomal compartment where it colocalizes with itch
Manually annotated by BRENDA team
the isoform Smurf1 C2 domain exerts a key role in localization to the plasma membrane. Lysine residues, Lys-28 and Lys-85, within the C2 domain are important for Smurf1 localization at the plasma membrane
Manually annotated by BRENDA team
mitochondrion-associated protein
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
physiological function
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
NEDD4_HUMAN
1319
0
149114
Swiss-Prot
Mitochondrion (Reliability: 5)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
190000
-
gel filtration
200000
x * 200000, SDS-PAGE
290000
-
dynamic light scattering
330000
-
dynamic light scattering, reoligomnerization of 80-kDa species
42200
x * 42200, enzyme including an additional five-residue peptide (Gly-Pro-Leu-Gly-Ser), calculated from amino acid sequence
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
trimer
-
3 * 80000, gel filtration
additional information
-
the predicted intersubunit interface has an absolutely conserved residue Phe823, substitution of which destabilizes the trimer and results in an over 10000fold decrease in kcat for polyubiquitin chain assembly
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ubiquitination
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
structure of ubiquitin-loaded human neural precursor cell–expressed developmentally downregulated protein, Nedd4, to 2.51 A resolution. The Nedd4-HECT domain-ubiquitin transitory intermediate provides a structural basis for the proposed sequential addition mechanism. The donor ubiquitin, transferred from the E2, is bound to the Nedd4 C lobe with its C-terminal tail locked in an extended conformation, primed for catalysis. Nedd4-family members are Lys63-specific enzymes whose catalysis is mediated by an essential C-terminal acidic residue
structures of Nedd4 HECT domain, alone (2.5 A) and in complex with ubiquitin (2.7 A), showing new binding modes involving two surfaces on ubiquitin and both subdomains of the HECT N-lobes, suggesting an model for the HECT-t-substrate ubiquitin transfer, in which the growing chain on the substrate is kept close to the catalytic cysteine to promote processivity HECTNedd4 displays the typical HECT fold, composed of two lobes connected by a flexible hinge. The N-lobe consists of two moieties, the large and the smal subdomains. The small domains host the E2-binding site and the large carries the catalytic cysteine
C2 domain, to 1.96 A resolution. The Smurf1 C2 domain possesses a typical anti-parallel beta-sandwich fold. The Smurf1 C2 domain exerts a key role in localization to the plasma membrane. Lysine residues, Lys-28 and Lys-85, within the C2 domain are important for Smurf1 localization at the plasma membrane, regulation on cell migration, and robust ligase activity toward RhoA, which further supports a Ca2+-independent localization mechanism for Smurf1
crystal structure of the extended HECT domain of AREL1 (amino acids 436-823) at 2.4 A resolution. The extended HECT domain adopts an inverted, T-shaped, bilobed conformation and harbors an additional loop (aa 567-573) absent in other HECT members. The N-terminal extended region (aa 436-482) preceding the HECT domain is indispensable for its stability and activity and without this region, the HECT domain becomes inactive
crystal structure of the HECT domain of human ubiquitin ligase WWP1/AIP5 maintains a two-lobed structure like the HECT domain of the human ubiquitin ligase E6AP. The organization of the two lobes relative one another is different from E6AP due to a rotation about a polypeptide hinge linking the N and C lobes
crystal structure of the HECT domain of UBE3C (amino acids 744-1083) with an additional fifty N-terminal amino acids (aa 693-743) at 2.7 A. The UBE3C HECT domain forms an open, L-shaped, bilobed conformation, having a large N-lobe and a small C-lobe. The N-terminal region (aa 693-743) preceding the UBE3C HECT domain as well as a loop region (aa 758-762) in the N-lobe of the HECT domain affect the stability and activity of UBE3C HECT domain
hanging drop vapor diffusion method, using 0.1 M HEPES pH 8.4, 0.2 M MgCl2, 15% (v/v) ethanol at 4°C
Huwe1 C-lobe-ubiquitin crystal structure at 2.8 A resolution, by molecular replacement and solution NMR spectroscopy
Smurf2 C-lobe-ubiquitin crystal structure at 2.8 A resolution, by molecular replacement and solution NMR spectroscopy
structures of ubiquitin variants in complex with the HECT domains of human E3 HECT ligases
-
structures of WWP1 in its fully inactive and partially active states. Domains WW2, L, and WW4 are organized into a headset architecture, in which the WW2 and WW4 domains are bound to bilateral sites within the N-lobe, and L forms a kinked alpha-helix that is tucked into the cleft between the N- and C-lobes of HECT
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
F707A
Nedd4 mutant, almost abolished HECTNedd4 binding to Lys 63 ubiquitin. Mutant F707A has defective chain elongation on substrate or shorter free chains
Y605A
Nedd4 mutant, almost abolished HECTNedd4 binding to Lys 63 ubiquitin
A349P
mutant is defective in thioester formation
A713P
mutant is defective in thioester formation
C1036A
loss of ubiquitylation activity
C2579G
-
mutation in catalytic cysteine, loss of E3 ligase activity
C716A
-
mutation in catalytic cysteine. Cells transfected with C716A fail to accumulate securin with significant reduction in Mad2 level. Cell show a significant increase in cells displaying misaligned chromosomes, lagging chromosomes during mitotic exit, and multinucleation
C922A
-
dominant-negative active site mutant, predicted to result in an end point of 33% activity compared with wild-type. Addition of mutant C922A to the wild-type enzyme at saturating concentration inhibits wild-type NEDD4L to 30% limiting activity
DELTA382-385
HECT domain mutant, capable of forming DTT-sensitive HECT-ubiquitin thioesters, defective in polyubiquitination
DELTA383-385
HECT domain mutant, capable of forming DTT-sensitive HECT-ubiquitin thioesters, defective in polyubiquitination
DELTA384-385
HECT domain mutant, capable of forming DTT-sensitive HECT-ubiquitin thioesters, defective in polyubiquitination
E411Q
slight decrease of affinity to human papilloma virus E6 oncogens
E411Q/E415Q
about 6fold decrease of affinity to human papilloma virus E6 oncogens
E415Q
slight decrease of affinity to human papilloma virus E6 oncogens
E415R
slight decrease of affinity to human papilloma virus E6 oncogens
E646A
-
400fold reduction in kcat value for polyubiquitination
E646D
-
1200fold reduction in kcat value for polyubiquitination
E701A
substitution in the Arel1 HECT domain, substantially increases its autopolyubiquitination and SMAC ubiquitination activity
E748A
mutant produces thioester levels comparable to the wild-type and shows enhanced polyubiquitination activity
F823A
-
10000fold reduction in kcat value
F823D
-
no detectable free or anchored polyubiquitin chain assembly is observed with the mutant
G383A/L384A
mutant produces thioester levels comparable to the wild-type
K1013Q
mutation increases the formation of E3-Ub thioester intermediate and shows 2fold enhanced ubiquitination activity
L346P
mutation abolishes E2-E3 transthiolation and consequently polyubiquitination activity
L384A
mutant produces thioester levels comparable to the wild-type
L409V
about 30fold decrease of affinity to human papilloma virus E6 oncogens characterised by very fast dissociation rates
L412V
moderate decrease of affinity to human papilloma virus E6 oncogens
L413S
about 40fold decrease of affinity to human papilloma virus E6 oncogens characterised by very fast dissociation rates
L413V
moderate decrease of affinity to human papilloma virus E6 oncogens
medicine
Q410E
no decrease of affinity to human papilloma virus E6 oncogens
Q961A
mutation of residues in the HECT domain, substantial decrease in autoubiquitination activity and in the formation of E3-Ub thioester intermediate
Q961E
mutation substantially reduces the formation of E3-Ub thioester intermediate
Q961E/K1013Q
mutant shows a reduced level of autoubiquitination activity
R604D
-
15fold reduction in kcat value for polyubiquitination
Ser1049
mutation of residues in the HECT domain, substantial decrease in autoubiquitination activity
V747A/E748A
mutant produces thioester levels comparable to the wild-type
additional information
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
34.5
melting temperature, HECT domain lacking loop residues 758-762
41
melting temperature, HECT domain
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Ni-NTA column chromatography and Superdex 200 gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
codon-optimized expression in Escherichia coli
-
expressed in Escherichia coli BL21(DE3) cells
expressed in HEK-293 cells
expression in Escherichia coli
expression in Escherihia coli
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
medicine
synthesis
construcution of a soluble HECT domain truncation of isoform WWP2 which is amendable for preparation scale expression in Escherichia coli. A relatively simple purification process achieves highly pure protein by employing immobilized metal-affinity chromatography followed by salting out, ion exchange chromatography and finally, size exclusion chromatography. Procedure allows to obtain about 60 mg/L of the soluble protein
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Nuber, U.; Scheffner, M.
Identification of determinants in E2 ubiquitin-conjugating enzymes required for hect E3 ubiquitin-protein ligase interaction
J. Biol. Chem.
274
7576-7582
1999
Saccharomyces cerevisiae (P39940), Saccharomyces cerevisiae, Homo sapiens (Q05086), Homo sapiens, Saccharomyces cerevisiae ATCC 204508 (P39940)
Manually annotated by BRENDA team
Yu, J.; Lan, J.; Zhu, Y.; Li, X.; Lai, X.; Xue, Y.; Jin, C.; Huang, H.
The E3 ubiquitin ligase HECTD3 regulates ubiquitination and degradation of Tara
Biochem. Biophys. Res. Commun.
367
805-812
2008
Homo sapiens (Q5T447)
Manually annotated by BRENDA team
Park, Y.; Yoon, S.K.; Yoon, J.B.
TRIP12 functions as an E3 ubiquitin ligase of APP-BP1
Biochem. Biophys. Res. Commun.
374
294-298
2008
Homo sapiens (Q14669)
Manually annotated by BRENDA team
Li, Y.; Ozaki, T.; Kikuchi, H.; Yamamoto, H.; Ohira, M.; Nakagawara, A.
A novel HECT-type E3 ubiquitin protein ligase NEDL1 enhances the p53-mediated apoptotic cell death in its catalytic activity-independent manner
Oncogene
27
3700-3709
2008
Homo sapiens (Q76N89)
Manually annotated by BRENDA team
Chen, A.; Gao, B.; Zhang, J.; McEwen, T.; Ye, S.Q.; Zhang, D.; Fang, D.
The HECT-type E3 ubiquitin ligase AIP2 inhibits activation-induced T-cell death by catalyzing EGR2 ubiquitination
Mol. Cell. Biol.
29
5348-5356
2009
Homo sapiens (O00308)
Manually annotated by BRENDA team
Lu, K.; Li, P.; Zhang, M.; Xing, G.; Li, X.; Zhou, W.; Bartlam, M.; Zhang, L.; Rao, Z.; He, F.
Pivotal role of the C2 domain of the Smurf1 ubiquitin ligase in substrate selection
J. Biol. Chem.
286
16861-16870
2011
Homo sapiens (Q9HCE7)
Manually annotated by BRENDA team
Maspero, E.; Mari, S.; Valentini, E.; Musacchio, A.; Fish, A.; Pasqualato, S.; Polo S.
Structure of the HECT:ubiquitin complex and its role in ubiquitin chain elongation
EMBO Rep.
12
342-349
2011
Homo sapiens (P46934)
Manually annotated by BRENDA team
Zanier, K.; Charbonnier, S.; Baltzinger, M.; Nomine, Y.; Altschuh, D.; Trave, G.
Kinetic analysis of the interactions of human papillomavirus E6 oncoproteins with the ubiquitin ligase E6AP using surface plasmon resonance
J. Mol. Biol.
349
401-412
2005
Homo sapiens (Q05086)
Manually annotated by BRENDA team
Lin, Q.; Wang, J.; Childress, C.; Sudol, M.; Carey, D.; Yang, W.
HECT E3 ubiquitin ligase Nedd4-1 ubiquitinates ACK and regulates epidermal growth factor (EGF)-induced degradation of EGF receptor and ACK
Mol. Cell. Biol.
30
1541-1554
2010
Homo sapiens (P46934)
Manually annotated by BRENDA team
Maspero, E.; Valentini, E.; Mari, S.; Cecatiello, V.; Soffientini, P.; Pasqualato, S.; Polo, S.
Structure of a ubiquitin-loaded HECT ligase reveals the molecular basis for catalytic priming
Nat. Struct. Mol. Biol.
20
696-701
2013
Homo sapiens (P46934), Homo sapiens
Manually annotated by BRENDA team
Li, Y.; Kong, Y.; Zhou, Z.; Chen, H.; Wang, Z.; Hsieh, Y.C.; Zhao, D.; Zhi, X.; Huang, J.; Zhang, J.; Li, H.; Chen, C.
The HECTD3 E3 ubiquitin ligase facilitates cancer cell survival by promoting K63-linked polyubiquitination of caspase-8
Cell Death Dis.
4
e935
2013
Homo sapiens (Q5T447)
Manually annotated by BRENDA team
Rossi, M.; Rotblat, B.; Ansell, K.; Amelio, I.; Caraglia, M.; Misso, G.; Bernassola, F.; Cavasotto, C.; Knight, R.; Ciechanover, A.; Melino, G.
High throughput screening for inhibitors of the HECT ubiquitin E3 ligase ITCH identifies antidepressant drugs as regulators of autophagy
Cell Death Dis.
5
e935
2014
Homo sapiens (Q96J02)
-
Manually annotated by BRENDA team
Nie, J.; Liu, L.; Wu, M.; Xing, G.; He, S.; Yin, Y.; Tian, C.; He, F.; Zhang, L.
HECT ubiquitin ligase Smurf1 targets the tumor suppressor ING2 for ubiquitination and degradation
FEBS Lett.
584
3005-3012
2010
Homo sapiens (Q9HCE7)
Manually annotated by BRENDA team
Angers, A.; Ramjaun, A.R.; McPherson, P.S.
The HECT domain ligase Itch ubiquitinates endophilin and localizes to the trans-Golgi network and endosomal system
J. Biol. Chem.
279
11471-11479
2004
Homo sapiens (Q96J02)
Manually annotated by BRENDA team
Park, Y.; Yoon, S.K.; Yoon, J.B.
The HECT domain of TRIP12 ubiquitinates substrates of the ubiquitin fusion degradation pathway
J. Biol. Chem.
284
1540-1549
2009
Homo sapiens (Q14669), Homo sapiens
Manually annotated by BRENDA team
Edwin, F.; Anderson, K.; Patel, T.B.
HECT domain-containing E3 ubiquitin ligase Nedd4 interacts with and ubiquitinates Sprouty2
J. Biol. Chem.
285
255-264
2010
Homo sapiens (P46934)
Manually annotated by BRENDA team
Tran, H.; Bustos, D.; Yeh, R.; Rubinfeld, B.; Lam, C.; Shriver, S.; Zilberleyb, I.; Lee, M.W.; Phu, L.; Sarkar, A.A.; Zohn, I.E.; Wertz, I.E.; Kirkpatrick, D.S.; Polakis, P.
HectD1 E3 ligase modifies adenomatous polyposis coli (APC) with polyubiquitin to promote the APC-axin interaction
J. Biol. Chem.
288
3753-3767
2013
Homo sapiens
Manually annotated by BRENDA team
Ding, Y.; Zhang, Y.; Xu, C.; Tao, Q.H.; Chen, Y.G.
HECT domain-containing E3 ubiquitin ligase NEDD4L negatively regulates Wnt signaling by targeting dishevelled for proteasomal degradation
J. Biol. Chem.
288
8289-8298
2013
Homo sapiens (Q96PU5)
Manually annotated by BRENDA team
Osmundson, E.C.; Ray, D.; Moore, F.E.; Gao, Q.; Thomsen, G.H.; Kiyokawa, H.
The HECT E3 ligase Smurf2 is required for Mad2-dependent spindle assembly checkpoint
J. Cell Biol.
183
267-277
2008
Homo sapiens
Manually annotated by BRENDA team
Verdecia, M.A.; Joazeiro, C.A.; Wells, N.J.; Ferrer, J.L.; Bowman, M.E.; Hunter, T.; Noel, J.P.
Conformational flexibility underlies ubiquitin ligation mediated by the WWP1 HECT domain E3 ligase
Mol. Cell
11
249-259
2003
Homo sapiens (Q9H0M0)
Manually annotated by BRENDA team
Jiang, J.; Zheng, J.; She, Y.; Jia, Z.
Expression and purification of human WWP2 HECT domain in Escherichia coli
Protein Expr. Purif.
110
95-101
2015
Homo sapiens (O00308)
Manually annotated by BRENDA team
Gong, W.; Zhang, X.; Zhang, W.; Li, J.; Li, Z.
Structure of the HECT domain of human WWP2
Acta Crystallogr. Sect. F
71
1251-1257
2015
Homo sapiens (O00308), Homo sapiens
Manually annotated by BRENDA team
An, C.I.; Ganio, E.; Hagiwara, N.
Trip12, a HECT domain E3 ubiquitin ligase, targets Sox6 for proteasomal degradation and affects fiber type-specific gene expression in muscle cells
Skeletal Muscle
3
11
2013
Homo sapiens (Q14669), Mus musculus
Manually annotated by BRENDA team
Singh, S.; Sivaraman, J.
Crystal structure of HECT domain of UBE3C E3 ligase and its ubiquitination activity
Biochem. J.
477
905-923
2020
Homo sapiens (Q15386)
Manually annotated by BRENDA team
Xu, Y.; Anderson, D.; Ye, Y.
The HECT domain ubiquitin ligase HUWE1 targets unassembled soluble proteins for degradation
Cell Discov.
2
16040
2016
Homo sapiens (Q7Z6Z7)
Manually annotated by BRENDA team
Byrne, R.; Mund, T.; Licchesi, J.
Activity-based probes for HECT E3 ubiquitin ligases
ChemBioChem
18
1415-1427
2017
Chlorocebus aethiops, Homo sapiens (Q15386)
Manually annotated by BRENDA team
Liu, Y.; HuangFu, W.; Huang, H.; Wu, W.; Chen, Y.; Yen, Y.; Huang, H.; Nien, C.; Lai, M.; Pan, S.; Liou, J.
1,4-Naphthoquinones as inhibitors of Itch, a HECT domain-E3 ligase, and tumor growth suppressors in multiple myeloma
Eur. J. Med. Chem.
140
84-91
2017
Homo sapiens (Q96J02)
Manually annotated by BRENDA team
Zhu, B.; Das, S.; Mitra, S.; Farris, T.; McBride, J.
Ehrlichia chaffeensis TRP120 moonlights as a HECT E3 ligase involved in selfand host ubiquitination to influence protein interactions and stability for intracellular survival
Infect. Immun.
85
e00290
2017
Ehrlichia chaffeensis, Homo sapiens (Q96PU5)
Manually annotated by BRENDA team
Braganza, A.; Li, J.; Zeng, X.; Yates, N.; Dey, N.; Andrews, J.; Clark, J.; Zamani, L.; Wang, X.; St Croix, C.; OSullivan, R.; Garcia-Exposito, L.; Brodsky, J.; Sobol, R.
UBE3B is a calmodulin-regulated, mitochondrion-associated E3 ubiquitin ligase
J. Biol. Chem.
292
2470-2484
2017
Homo sapiens (Q7Z3V4), Homo sapiens
Manually annotated by BRENDA team
Todaro, D.R.; Augustus-Wallace, A.C.; Klein, J.M.; Haas, A.L.
Oligomerization of the HECT ubiquitin ligase NEDD4-2/NEDD4L is essential for polyubiquitin chain assembly
J. Biol. Chem.
293
18192-18206
2018
Homo sapiens
Manually annotated by BRENDA team
Mund, T.; Pelham, H.
Substrate clustering potently regulates the activity of WW-HECT domain-containing ubiquitin ligases
J. Biol. Chem.
293
5200-5209
2018
Homo sapiens (O00308)
Manually annotated by BRENDA team
Singh, S.; Ng, J.; Nayak, D.; Sivaraman, J.
Structural insights into a HECT-type E3 ligase AREL1 and its ubiquitination activities in vitro
J. Biol. Chem.
294
19934-19949
2019
Homo sapiens (O15033)
Manually annotated by BRENDA team
Lin, Q.; Dai, Q.; Meng, H.; Sun, A.; Wei, J.; Peng, K.; Childress, C.; Chen, M.; Shao, G.; Yang, W.
The HECT E3 ubiquitin ligase NEDD4interacts with and ubiquitylates SQSTM1 for inclusion body autophagy
J. Cell Sci.
130
3839-3850
2017
Homo sapiens (P46934)
Manually annotated by BRENDA team
Jaeckl, M.; Stollmaier, C.; Strohaeker, T.; Hyz, K.; Maspero, E.; Polo, S.; Wiesner, S.
beta-Sheet augmentation is a conserved mechanism of priming HECT E3 ligases for ubiquitiniquitin ligation
J. Mol. Biol.
430
3218-3233
2018
Homo sapiens (Q7Z6Z7), Homo sapiens (Q9HAU4)
Manually annotated by BRENDA team
Zhang, W.; Wu, K.; Sartori, M.; Kamadurai, H.; Ordureau, A.; Jiang, C.; Mercredi, P.; Murchie, R.; Hu, J.; Persaud, A.; Mukherjee, M.; Li, N.; Doye, A.; Walker, J.; Sheng, Y.; Hao, Z.; Li, Y.; Brown, K.; Lemichez, E.; Chen, J.; Tong, Y.; Harper, J.; Moffat, J.
System-wide modulation of HECT E3 ligases with selective ubiquitin variant probes
Mol. Cell.
62
121-136
2016
Homo sapiens
Manually annotated by BRENDA team
Wang, Z.; Liu, Z.; Chen, X.; Li, J.; Yao, W.; Huang, S.; Gu, A.; Lei, Q.; Mao, Y.; Wen, W.
A multi-lock inhibitory mechanism for fine-tuning enzyme activities of the HECT family E3 ligases
Nat. Commun.
10
3162
2019
Homo sapiens (Q9H0M0), Homo sapiens
Manually annotated by BRENDA team