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PXD021393

PXD021393 is an original dataset announced via ProteomeXchange.

Dataset Summary
TitleMultivalent lipid targeting by the calcium-independent C2A domain of Slp-4/granuphilin
DescriptionSynaptotagmin-like protein 4 (Slp-4), also known as granuphilin, is a Rab effector responsible for docking insulin secretory vesicles to the plasma membrane prior to exocytosis. Slp-4 binds vesicular Rab proteins via an N-terminal Slp homology (SHD) domain, interacts with plasma membrane SNARE complex proteins via a central linker region, and contains tandem C-terminal C2 domains (C2A and C2B) with affinity for phosphatidylinositol-(4,5)-bisphosphate (PIP2). Its C2A domain has previously been shown to bind PIP2 or its soluble analogues with low micromolar affinity; however, the domain docks with low nanomolar apparent affinity to PIP2 in lipid vesicles that also contain background anionic lipids such as phosphatidylserine (PS). Here we show using a combination of computational and experimental approaches that this high-affinity membrane interaction arises from concerted interaction at multiple sites on the C2A domain. In addition to the previously identified, PIP2-selective lysine cluster, a larger cationic surface surrounding the cluster contributes substantially to the affinity for physiologically relevant lipid compositions. While mutations at the PIP2-selective site decreases affinity for PIP2, multiple mutations are needed to decrease binding to physiologically relevant lipid compositions. Docking and molecular dynamics simulations indicate several conformationally flexible loops that contribute to the nonspecific cationic surface. In addition, we identify and characterize a covalently modified variant in the bacterially expressed protein, which arises through reactivity of the PIP2-binding lysine cluster with endogenous bacterial compounds and has a low membrane affinity. Overall, multivalent lipid binding by the Slp-4 C2A domain provides selective recognition and high affinity docking of large dense-core secretory vesicles to the plasma membrane.
HostingRepositoryPRIDE
AnnounceDate2024-10-08
AnnouncementXMLSubmission_2024-10-08_11:18:46.693.xml
DigitalObjectIdentifier
ReviewLevelPeer-reviewed dataset
DatasetOriginOriginal dataset
RepositorySupportUnsupported dataset by repository
PrimarySubmitterCole Michel
SpeciesList scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationListNo PTMs are included in the dataset
Instrument6520A Quadrupole Time-of-Flight LC/MS
Dataset History
RevisionDatetimeStatusChangeLog Entry
02020-09-10 01:38:09ID requested
12024-10-08 11:18:47announced
Publication List
10.1074/jbc.ra120.014618;
Alnaas AA, Watson-Siriboe A, Tran S, Negussie M, Henderson JA, Osterberg JR, Chon NL, Harrott BM, Oviedo J, Lyakhova T, Michel C, Reisdorph N, Reisdorph R, Shearn CT, Lin H, Knight JD, Multivalent lipid targeting by the calcium-independent C2A domain of synaptotagmin-like protein 4/granuphilin. J Biol Chem, 296():100159(2021) [pubmed]
Keyword List
submitter keyword: granuphilin,Intact Protein, phosphogluconate, LCMS
Contact List
Cole Robert Michel
contact affiliationResidorph Lab, Mass Spectrometry Core - Skaggs School of Pharmacy, University of Colorado - Anschutz Medical Campus
contact emailcole.michel@cuanschutz.edu
lab head
Cole Michel
contact affiliationUniveristy of Colorado - Skaggs School of Pharmacy
contact emailcole.michel@cuanschutz.edu
dataset submitter
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