PXD018219

PXD018219 is an original dataset announced via ProteomeXchange.

Title	Inter- and Intra-Molecular Interactions of The Arabidopsis Plasma Membrane Proton Pump Revealed Using a Mass Spectrometric-Cleavable Crosslinker
Description	In plants and fungi, the plasma membrane proton pump (H+-ATPase) establishes an electrochemical gradient across the plasma membrane which serves as the driving force for the secondary transport of ions and nutrients across the cell membrane. This is an essential enzyme that functions in many important processes including stomatal movement, cell elongation, and cellular responses to stimuli from hormones, light, and other environmental conditions. Therefore, understanding how the activity of the H+-ATPase is regulated is important to understand how plants adapt to different growth conditions. The autoinhibitory effect of the C- terminal regulatory domain of H+-ATPase is well established and is thought to be mediated by interactions with the catalytic domains. Here, using the lysine reactive mass spectrometry cleavable crosslinker DSSO, we found that the C-terminal domain of the Arabidopsis H+- ATPase 2 (AHA2) crosslinked extensively with the actuator, nucleotide-binding, and phosphorylation domains, suggesting that the C-terminal domain regulates the catalytic cycle by modulating the relative positions of these domains. Interestingly, several C-terminal crosslinks occurred near a predicted proton binding site (Asp-684 in TM6), suggesting that the C-terminal domain may regulate proton efflux. Additionally, crosslinks between the C-terminal domain and other domains of AHA2 were detected in a monomeric protein resolved on SDS-PAGE, suggesting that intramolecular interactions may also be involved in the regulation of enzyme activity. Finally, we observed mixed-isotope crosslinking between unlabeled (14N-AHA2) and labeled (15N-AHA2) between the C-terminal domain and other domains of AHA2, supporting our model that oligomeric H+-ATPase may autoinhibit the neighboring monomer in a “head to tail” configuration.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_08:48:26.013.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Thao Thi Nguyen
SpeciesList	scientific name: Arabidopsis thaliana (Mouse-ear cress); NCBI TaxID: 3702; scientific name: Saccharomyces cerevisiae (Baker's yeast); NCBI TaxID: 4932;
ModificationList	monohydroxylated residue; iodoacetamide derivatized residue; deamidated residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2020-03-26 15:17:58	ID requested
1	2023-06-01 11:49:54	announced
⏵ 2	2023-11-14 08:48:29	announced	2023-11-14: Updated project metadata.

Nguyen TT, Blackburn MR, Sussman MR, Plasma Membrane Proton Pump Revealed Using a Mass Spectrometry Cleavable Cross-Linker. Biochemistry, 59(24):2210-2225(2020) [pubmed]

submitter keyword: Arabidopsis, plasma membrane protein pump, metabolic labeling,DSSO, crosslinking

Michael R. Sussman
contact affiliation	University of Wisconsin Madison, Department of Biochemistry
contact email	msussman@wisc.edu
lab head
Thao Thi Nguyen
contact affiliation	University of Missouri-Columbia
contact email	tnguyen24@wisc.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD018219
     1. Label: PRIDE project
     2. Name: Inter- and Intra-Molecular Interactions of The Arabidopsis Plasma Membrane Proton Pump Revealed Using a Mass Spectrometric-Cleavable Crosslinker