PXD045837

PXD045837 is an original dataset announced via ProteomeXchange.

Title	Structural mechanisms of autoinhibition and substrate recognition by the ubiquitin ligase HACE1
Description	Ubiquitin ligases (E3s) are pivotal specificity determinants in the ubiquitin system by selecting substrates and decorating them with distinct ubiquitin signals. Structure determination of the underlying, specific E3-substrate complexes, however, has proven challenging due to their transient nature. In particular, it is incompletely understood how members of the catalytic cysteine-driven class of HECT-type ligases position substrate proteins for modification. Here we report a cryo-EM structure of the full-length human HECT-type ligase HACE1, along with solution-based conformational analyses by small-angle X-ray scattering and hydrogen-deuterium exchange mass spectrometry. Structure-based functional analyses in vitro and in cells reveal that the activity of HACE1 is stringently regulated by dimerization-induced autoinhibition. The inhibition occurs at the first step of the catalytic cycle and is thus substrate-independent. We employ mechanism-based chemical crosslinking to reconstitute a complex of activated, monomeric HACE1 with its major substrate, RAC1, visualize its structure by cryo-EM, and validate the binding mode by solution-based analyses. Our findings explain how HACE1 achieves selectivity in ubiquitinating the active, GTP-loaded state of RAC1 and establish a framework for interpreting mutational alterations of the HACE1-RAC1 interplay in disease. More broadly, this work illuminates central unexplored aspects in the architecture, conformational dynamics, regulation, and specificity of full-length HECT-type ligases.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_06:29:13.981.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Wieland Steinchen
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Synapt MS

Revision	Datetime	Status	ChangeLog Entry
0	2023-09-30 05:06:30	ID requested
1	2024-02-12 12:41:09	announced
⏵ 2	2024-10-22 06:29:14	announced	2024-10-22: Updated project metadata.

10.1038/S41594-023-01203-4;

ProteomeXchange project tag: Hydrogen Deuterium Exchange (HDX-MS)

submitter keyword: HDX-MS

Wieland Steinchen
contact affiliation	Philipps-Universität Marburg Zentrum für Synthetische Mikrobiologie (SYNMIKRO) Karl-von-Frisch-Straße 14 35043 Marburg
contact email	wieland.steinchen@synmikro.uni-marburg.de
lab head
Wieland Steinchen
contact affiliation	Philipps-University Marburg, Department of Chemistry & SYNMIKRO, Karl-von-Frisch-Straße 14, 35043 Marburg, GERMANY
contact email	wieland.steinchen@synmikro.uni-marburg.de
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2024/02/PXD045837

PRIDE project URI

• PRIDE
  1. PXD045837
     1. Label: PRIDE project
     2. Name: Structural mechanisms of autoinhibition and substrate recognition by the ubiquitin ligase HACE1