PXD031606

PXD031606 is an original dataset announced via ProteomeXchange.

Title	Sequence and structural Sequence and structural variations determining the recruitment of WNK kinases to the KLHL3 E3 ligase
Description	The BTB-Kelch protein KLHL3 is a Cullin3-dependent E3 ligase that mediates the ubiquitin-dependent degradation of kinases WNK1-4 to control blood pressure and cell volume. A crystal structure of KLHL3 has defined its binding to an acidic degron motif containing a PXXP sequence that is strictly conserved in WNK1, WNK2 and WNK4. Mutations in the second proline abrograte the interaction causing the hypertension syndrome pseudohypoaldosteronism type II. WNK3 shows a diverged degron motif containing 4 amino acid substitutions that remove the PXXP motif raising questions as to the mechanism of its binding. To understand this atypical interaction, we determined the crystal structure of the KLHL3 Kelch domain in complex with a WNK3 peptide. The electron density enabled the complete 11-mer WNK-family degron motif to be traced for the first time revealing several conserved features not captured in previous work, including additional salt bridge and hydrogen bond interactions. Overall, the WNK3 peptide adopted a conserved binding pose except for a subtle shift to accommodate bulkier amino acid substitutions at the binding interface. At the centre, the second proline was substituted by WNK3 Thr541, providing a unique phosphorylatable residue among the WNK-family degrons. Fluorescence polarisation and structural modelling experiments revealed that its phosphorylation would abrogate the KLHL3 interaction similarly to hypertension-causing mutations. Together, these data reveal how the KLHL3 Kelch domain can accommodate the binding of multiple WNK isoforms and highlight a potential regulatory mechanism for the recruitment of WNK3.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_08:55:19.948.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD031606
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Roman Fischer
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	phosphorylated residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2022-02-11 06:16:05	ID requested
1	2022-03-07 06:11:43	announced
⏵ 2	2023-11-14 08:55:20	announced	2023-11-14: Updated project metadata.

10.6019/PXD031606;

submitter keyword: KLHL3-WNK3 complex,Structure

Roman Fischer
contact affiliation	Discovery Proteomics Facility Target Discovery Institute University of Oxford
contact email	roman.fischer@ndm.ox.ac.uk
lab head
Roman Fischer
contact affiliation	University of Oxford
contact email	roman.fischer@ndm.ox.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD031606
     1. Label: PRIDE project
     2. Name: Sequence and structural Sequence and structural variations determining the recruitment of WNK kinases to the KLHL3 E3 ligase