Dataset Information

Photo-affinity labeling with photo-sanglifehrin A probes

ABSTRACT: The interactome of 2 photo-Sanglifehrin A (pSfA) probes in Jurkat and K562 cells was profiled.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Christina Woo

LAB HEAD: Christina M. Woo

PROVIDER: PXD029540 | Pride | 2024-10-17

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	0419302619LHF_SAM05419_Hope389_HpH1.raw	Raw
	0419302619LHF_SAM05419_Hope389_HpH1_20190502001118.raw	Raw
	0419302619LHF_SAM05419_Hope389_HpH2.raw	Raw
	0419302619LHF_SAM05419_Hope389_HpH4.raw	Raw
	0419302619LHF_SAM05419_Hope389_HpH5.raw	Raw

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Publications

Sanglifehrin A mitigates multiorgan fibrosis by targeting the collagen chaperone cyclophilin B.

Flaxman Hope A HA Chrysovergi Maria-Anna MA Han Hongwei H Kabir Farah F Lister Rachael T RT Chang Chia-Fu CF Yvon Robert R Black Katharine E KE Weigert Andreas A Savai Rajkumar R Egea-Zorrilla Alejandro A Pardo-Saganta Ana A Lagares David D Woo Christina M CM

JCI insight 20240620 15

Pathological deposition and crosslinking of collagen type I by activated myofibroblasts drives progressive tissue fibrosis. Therapies that inhibit collagen synthesis have potential as antifibrotic agents. We identify the collagen chaperone cyclophilin B as a major cellular target of the natural product sanglifehrin A (SfA) using photoaffinity labeling and chemical proteomics. Mechanistically, SfA inhibits and induces the secretion of cyclophilin B from the endoplasmic reticulum (ER) and prevents ...[more]

PMID: 38900587

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Project description:PRIDE > Archive > PXD013420 Project PXD013420 Download Project Files Summary Title A binding site hotspot map of the FKBP12–rapamycin–FRB ternary complex by photo-affinity labeling and mass spectrometry-based proteomics Description Structural characterization of small molecule binding site hotspots within the global proteome is uniquely enabled by photo-affinity labeling (PAL) coupled with chemical enrichment and unbiased analysis by mass spectrometry (MS). MS-based binding site hotspot maps provide structural resolution of interaction sites in conjunction with identification of target proteins. However, binding site hotspot mapping has been confined to relatively simple small molecules to date; extension to more complex compounds would enable the structural definition of new binding modes in the proteome. Here, we extend PAL and MS methods to derive a binding site hotspot map for the immunosuppressant rapamycin, a complex macrocyclic natural product that forms a ternary complex with the proteins FKBP12 and FRB. Photo-rapamycin was developed as a diazirine-based PAL probe for rapamycin, and the FKBP12–photo-rapamycin–FRB ternary complex formed readily in vitro. Photo-irradiation, digestion, and MS analysis of the ternary complex revealed a McLafferty rearrangement product of photo-rapamycin conjugated to specific surfaces on FKBP12 and FRB. Molecular modeling of the ternary complex based on the binding site map revealed a 5.0 Å minimum distance constraint between the conjugated residues and the diazirine carbon. Molecular dynamics further predicted a 9.0 Å labeling radius for the diazirine upon photo-activation that may be useful in the interpretation of binding site measurements from PAL more broadly. Thus, in characterizing the ternary complex of photo-rapamycin by MS, we applied binding site hotspot mapping to a macrocyclic natural product and extracted a precise structural measurement for interpretation of PAL products that may enable the discovery of new ligand space in the “undruggable” proteome.