HECTc

Domain Homologous to E6-AP Carboxyl Terminus with

• SMART accession number: SM00119
• Description: E3 ubiquitin-protein ligases. Can bind to E2 enzymes.
• Interpro abstract (IPR000569):

  The HECT (Homologous to the E6-AP Carboxyl Terminus) domain is an around 350 amino acids motif that has been identified in proteins that all belong to a particular E3 ubiquitin-protein ligase family [ (PUBMED:7708685) ]. HECT domain containing proteins accept ubiquitin from an E2 ubiquitin-conjugating enzyme in the form of a thioester and then transfer it to lysine side chains of target proteins, and transfers additional ubiquitin molecules to the end of growing ubiquitin chains. The site of ubiquitin thioester formation is a conserved cysteine residue located in the last 32-36 aa of the HECT domain [ (PUBMED:7708685) ]. The amino-terminal part of the HECT domain has been involved in E2 binding [ (PUBMED:9182527) (PUBMED:9153201) ]. Once linked to ubiquitin, the target proteins are degraded in the 26 S proteasome.

• GO function: ubiquitin-protein transferase activity (GO:0004842)

There are 23694 HECTc domains in 23671 proteins in SMART's nrdb database.

Cellular role (predicted cellular role)

Binding / catalysis: ubiquitin-protein ligase

Literature (relevant references for this domain)

Primary literature is listed below; Automatically-derived, secondary literature is also avaliable.

• Schwarz SE, Rosa JL, Scheffner M
• Characterization of human hect domain family members and their interaction with UbcH5 and UbcH7.
• J Biol Chem. 1998; 273: 12148-54

The hect domain protein family was originally identified by sequence similarity of its members to the C-terminal region of E6-AP, an E3 ubiquitin-protein ligase. Since the C terminus of E6-AP mediates thioester complex formation with ubiquitin, a necessary intermediate step in E6-AP-dependent ubiquitination, it was proposed that members of the hect domain family in general have E3 activity. The hect domain is approximately 350 amino acids in length, and we show here that the hect domain of E6-AP is necessary and sufficient for ubiquitin thioester adduct formation. Furthermore, the human genome encodes at least 20 different hect domain proteins, and in further support of the hypothesis that hect domain proteins represent a family of E3s, several of these are shown to form thioester complexes with ubiquitin. In addition, some hect domain proteins interact preferentially with UbcH5, whereas others interact with UbcH7, indicating that human hect domain proteins can be grouped into at least two classes based on their E2 specificity. Since E3s are thought to play a major role in substrate recognition, the presence of a large family of E3s should contribute to ensure the specificity and selectivity of ubiquitin-dependent proteolytic pathways.

• Hatakeyama S, Jensen JP, Weissman AM
• Subcellular localization and ubiquitin-conjugating enzyme (E2) interactions of mammalian HECT family ubiquitin protein ligases.
• J Biol Chem. 1997; 272: 15085-92

In most instances, the transfer of ubiquitin to target proteins is catalyzed by the action of ubiquitin protein ligases (E3s). Full-length cDNAs encoding murine E6-associated protein (mE6-AP) as well as Nedd-4, a protein that is homologous to E6-AP in its C terminus, were cloned. Nedd-4 and mouse E6-AP are both enzymatically active E3s and function with members of the UbcH5 family of E2s. Mouse E6-AP, like its human counterpart, ubiquitinates p53 in the presence of human papilloma virus E6 protein, while Nedd-4 does not. Consistent with its role in p53 ubiquitination, mE6-AP was found both in the nucleus and cytosol, while Nedd-4 was found only in the cytosol. Binding studies implicate a 150-amino acid region that is 40% identical between mE6-AP and Nedd-4 as a binding site for the C-terminal portion of an E2 enzyme (UbcH5B). Nedd-4 was determined to have a second nonoverlapping E2 binding site that recognizes the first 67 amino acids of UbcH5B but not the more C-terminal portion of this E2. These findings provide the first demonstration of physical interactions between mammalian E2s and E3s and establish that these interactions occur independently of ubiquitin and an intact E3 catalytic domain. Furthermore, the presence of two E2 binding sites within Nedd-4 suggests models for ubiquitination involving multiple E2 enzymes associated with E3s.

• Kumar S, Kao WH, Howley PM
• Physical interaction between specific E2 and Hect E3 enzymes determines functional cooperativity.
• J Biol Chem. 1997; 272: 13548-54

The cellular protein E6AP functions as an E3 ubiquitin protein ligase in the E6-dependent ubiquitination of p53. E6AP is a member of a family of functionally related E3 proteins that share a conserved carboxyl-terminal region called the Hect domain. Although several different E2 ubiquitin-conjugating enzymes have been shown to function with E6AP in the E6-dependent ubiquitination of p53 in vitro, the E2s that cooperate with E6AP in the ubiquitination of its normal substrates are presently unknown. Moreover, the basis of functional cooperativity between specific E2 and Hect E3 proteins has not yet been determined. Here we report the cloning of a new human E2, designated UbcH8, that was identified in a two-hybrid screen through specific interaction with E6AP. We demonstrate that UbcH7, an E2 closely related to UbcH8, can also bind to E6AP. The region of E6AP involved in complex formation with UbcH8 and UbcH7 was mapped to its Hect domain. Furthermore, we show that UbcH5 and UbcH6, two highly homologous E2s that were deficient for interaction with E6AP, could associate efficiently with another Hect-E3 protein, RSP5. Finally, only the E6AP-interacting E2s could function in conjunction with E6AP in the ubiquitination of an E6 independent substrate of E6AP, whereas the noninteracting E2s could not. Taken together, these studies demonstrate for the first time complex formation between specific human E2s and the Hect domain family of E3 proteins and suggest that selective physical interaction between E2 and E3 enzymes forms the basis of specificity for functionally distinct E2:E3 combinations.

• Nuber U, Schwarz S, Kaiser P, Schneider R, Scheffner M
• Cloning of human ubiquitin-conjugating enzymes UbcH6 and UbcH7 (E2-F1) and characterization of their interaction with E6-AP and RSP5.
• J Biol Chem. 1996; 271: 2795-800

E6-AP, a 100-kDa cellular protein, was originally identified through its interaction with the E6 protein of the oncogenic human papillomavirus types 16 and 18. The complex of E6-AP and E6 specifically interacts with p53 and mediates ubiquitination of p53 in concert with the E1 ubiquitin-activating enzyme and the E2 ubiquitin-conjugating enzyme UbcH5. Recent results suggest that E6-AP is representative of a family of putative ubiquitin-protein ligases. Members of this family are characterized by a conserved C-terminal region, termed hect domain. In this paper, we describe the isolation of two human E2s, designated as UbcH6 and UbcH7, that in addition to UbcH5 can interact with E6-AP. UbcH6 is a novel member of an evolutionally conserved subfamily of E2s that includes UbcH5 and Saccharomyces cerevisiae UBC4. Although UbcH7 does not appear to be a member of this subfamily, UbcH7 efficiently substitutes for UbcH5 in E6-AP-dependent ubiquitination. Surprisingly, UbcH6 was only weakly active in this particular assay. In addition, UbcH5 but not UbcH6 or UbcH7 efficiently interacts with the heet protein RSP5. These results indicate that E6-AP can interact with at least two species of E2 and that different hect proteins may interact with different E2s.

• Huibregtse JM, Scheffner M, Beaudenon S, Howley PM
• A family of proteins structurally and functionally related to the E6-AP ubiquitin-protein ligase.
• Proc Natl Acad Sci U S A. 1995; 92: 2563-7

E6-AP is a 100-kDa cellular protein that interacts with the E6 protein of the cancer-associated human papillomavirus types 16 and 18. The E6/E6-AP complex binds to and targets the p53 tumor-suppressor protein for ubiquitin-mediated proteolysis. E6-AP is an E3 ubiquitin-protein ligase which accepts ubiquitin from an E2 ubiquitin-conjugating enzyme in the form of a thioester and then directly transfers the ubiquitin to targeted substrates. The amino acid sequence of E6-AP shows similarity to a number of protein sequences over an approximately 350-aa region corresponding to the carboxyl termini of both E6-AP and the E6-AP-related proteins. Of particular note is a conserved cysteine residue within the last 32-34 aa, which in E6-AP is likely to be the site of ubiquitin thioester formation. Two of the E6-AP-related proteins, a rat 100-kDa protein and a yeast 95-kDa protein (RSP5), both of previously unknown function, are shown here to form thioesters with ubiquitin. Mutation of the conserved cysteine residue of these proteins destroys their ability to accept ubiquitin. These data strongly suggest that the rat 100-kDa protein and RSP5, as well as the other E6-AP-related proteins, belong to a class of functionally related E3 ubiquitin-protein ligases, defined by a domain homologous to the E6-AP carboxyl terminus (hect domain).

Metabolism (metabolic pathways involving proteins which contain this domain)

% proteins involved	KEGG pathway ID	Description
35.29	map04120	Ubiquitin mediated proteolysis
29.41	map00380	Tryptophan metabolism
19.12	map05050	Dentatorubropallidoluysian atrophy (DRPLA)
16.18	map04350	TGF-beta signaling pathway

This information is based on mapping of SMART genomic protein database to KEGG orthologous groups. Percentage points are related to the number of proteins with HECTc domain which could be assigned to a KEGG orthologous group, and not all proteins containing HECTc domain. Please note that proteins can be included in multiple pathways, ie. the numbers above will not always add up to 100%.

Structure (3D structures containing this domain)

3D Structures of HECTc domains in PDB

PDB code	Title
1c4z	STRUCTURE OF E6AP: INSIGHTS INTO UBIQUITINATION PATHWAY
1d5f	STRUCTURE OF AN E6AP-UBCH7 COMPLEX: INSIGHTS INTO THE UBIQUITINATION PATHWAY
1nd7	Conformational Flexibility Underlies Ubiquitin Ligation Mediated by the WWP1 HECT domain E3 Ligase
1zvd	Regulation of Smurf2 Ubiquitin Ligase Activity by Anchoring the E2 to the HECT domain
2oni	Catalytic Domain of the Human NEDD4-like E3 Ligase
2xbb	Nedd4 HECT:Ub complex
2xbf	Nedd4 HECT structure
3g1n	Catalytic domain of the human E3 ubiquitin-protein ligase HUWE1
3h1d	Structure of the HUWE1 HECT Domain
3jvz	E2~Ubiquitin-HECT
3jw0	E2~Ubiquitin-HECT
3olm	Structure and Function of a Ubiquitin Binding Site within the Catalytic Domain of a HECT Ubiquitin Ligase
3pt3	Crystal structure of the C-terminal lobe of the human UBR5 HECT domain
3tug	Crystal structure of the HECT domain of ITCH E3 ubiquitin ligase
4bbn	NEDD4 HECT-Ub:Ub complex
4be8	NEDD4 HECT A889F structure
4lcd	Structure of an Rsp5xUbxSna3 complex: Mechanism of ubiquitin ligation and lysine prioritization by a HECT E3
4y07	4Y07
5c7j	5C7J
5c7m	5C7M
5c91	5C91
5hpk	5HPK
5hpl	5HPL
5hps	5HPS
5hpt	5HPT

Links (links to other resources describing this domain)

• INTERPRO: IPR000569