PDBsum entry 1fxt

Ligase

PDB id: 1fxt

Contents

Protein chains

149 a.a. *

76 a.a. *

* Residue conservation analysis

PDB id:

1fxt

Name:

Ligase

Title:

Structure of a conjugating enzyme-ubiquitin thiolester complex

Structure:

Ubiquitin-conjugating enzyme e2-24 kda. Chain: a. Engineered: yes. Ubiquitin. Chain: b. Engineered: yes

Source:

Saccharomyces cerevisiae. Baker's yeast. Organism_taxid: 4932. Expressed in: escherichia coli. Expression_system_taxid: 562. Homo sapiens. Human. Organism_taxid: 9606.

NMR struc:

1 models

Authors:

K.S.Hamilton,G.S.Shaw,R.S.Williams,J.T.Huzil,S.Mckenna,C.Ptak, M.Glover,M.J.Ellison

Key ref:

K.S.Hamilton et al. (2001). Structure of a conjugating enzyme-ubiquitin thiolester intermediate reveals a novel role for the ubiquitin tail. Structure, 9, 897-904. PubMed id: 11591345 DOI: 10.1016/S0969-2126(01)00657-8

Date:

26-Sep-00 Release date: 10-Oct-01

Protein chain

P21734  (UBC1_YEAST) -  Ubiquitin-conjugating enzyme E2 1 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 215 a.a.

Struc: 149 a.a.

Protein chain

P0CG48  (UBC_HUMAN) -  Polyubiquitin-C from Homo sapiens

Seq: 685 a.a.

Struc: 76 a.a.

Enzyme reactions

• Enzyme class 2: Chain A: E.C.2.3.2.23 - E2 ubiquitin-conjugating enzyme.
• Reaction: S-ubiquitinyl-[E1 ubiquitin-activating enzyme]-L-cysteine + [E2 ubiquitin-conjugating enzyme]-L-cysteine = [E1 ubiquitin-activating enzyme]-L-cysteine + S-ubiquitinyl-[E2 ubiquitin-conjugating enzyme]-L- cysteine
• Enzyme class 3: Chain B: E.C.?

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

DOI no: 10.1016/S0969-2126(01)00657-8

Structure 9:897-904 (2001)

PubMed id: 11591345

Structure of a conjugating enzyme-ubiquitin thiolester intermediate reveals a novel role for the ubiquitin tail.

K.S.Hamilton, M.J.Ellison, K.R.Barber, R.S.Williams, J.T.Huzil, S.McKenna, C.Ptak, M.Glover, G.S.Shaw.

ABSTRACT

BACKGROUND: Ubiquitin-conjugating enzymes (E2s) are central enzymes involved in ubiquitin-mediated protein degradation. During this process, ubiquitin (Ub) and the E2 protein form an unstable E2-Ub thiolester intermediate prior to the transfer of ubiquitin to an E3-ligase protein and the labeling of a substrate for degradation. A series of complex interactions occur among the target substrate, ubiquitin, E2, and E3 in order to efficiently facilitate the transfer of the ubiquitin molecule. However, due to the inherent instability of the E2-Ub thiolester, the structural details of this complex intermediate are not known. RESULTS: A three-dimensional model of the E2-Ub thiolester intermediate has been determined for the catalytic domain of the E2 protein Ubc1 (Ubc1(Delta450)) and ubiquitin from S. cerevisiae. The interface of the E2-Ub intermediate was determined by kinetically monitoring thiolester formation by 1H-(15)N HSQC spectra by using combinations of 15N-labeled and unlabeled Ubc1(Delta450) and Ub proteins. By using the surface interface as a guide and the X-ray structures of Ub and the 1.9 A structure of Ubc1(Delta450) determined here, docking simulations followed by energy minimization were used to produce the first model of a E2-Ub thiolester intermediate. CONCLUSIONS: Complementary surfaces were found on the E2 and Ub proteins whereby the C terminus of Ub wraps around the E2 protein terminating in the thiolester between C88 (Ubc1(Delta450)) and G76 (Ub). The model supports in vivo and in vitro experiments of E2 derivatives carrying surface residue substitutions. Furthermore, the model provides insights into the arrangement of Ub, E2, and E3 within a ternary targeting complex.

Selected figure(s)

Figure 3.
Figure 3. Model of the E2-Ub Thiolester IntermediateThe model was determined by Monte Carlo docking as described in Experimental Procedures. The model shows (a) side and (b) end orientation views of helix a2 in the E2 molecule. Residues are indicated on both E2 and Ub to indicate important side chain-side chain interactions that arise at the protein-protein interface, as described in the text. In both figures, the thiol-forming C88 residue in E2 is a shown as a yellow ball-and-stick representation near the (a) center and (b) top of the complex

The above figure is reprinted by permission from Cell Press: Structure (2001, 9, 897-904) copyright 2001.

Figure was selected by an automated process.