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Quantitative Analysis of Samples Labeled with Multiplex iqPIR Cross-Linkers

ABSTRACT: The isobaric quantitative protein interaction reporter (iqPIR) has now been extended to six channels, enabling quantitation of cross-links from six different samples simultaneously. To analyze data for such cross-linkers, the two-channel mode of quantitative analysis was extended to accommodate any number of channels with defined isotope peak offsets. The HeLa 1:1:1:1:1 data was used for the development of the quantitation method of the 6-plex iqPIR.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Hela Cell

DISEASE(S): Cervix Carcinoma

SUBMITTER: Helisa Wippel

LAB HEAD: James E. Bruce

PROVIDER: PXD025796 | Pride | 2023-03-11

REPOSITORIES: Pride

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			Action	DRS
	022721_HeLa_6plex_1_1_F10_mango_2rep_1.raw	Raw
	022721_HeLa_6plex_1_1_F10_mango_2rep_2.raw	Raw
	022721_HeLa_6plex_1_1_F11_14_mango_2rep_1.raw	Raw
	022721_HeLa_6plex_1_1_F11_14_mango_2rep_2.raw	Raw
	022721_HeLa_6plex_1_1_F6_7_mango_2rep_1.raw	Raw

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Publications

Multiplexed Cross-Linking with Isobaric Quantitative Protein Interaction Reporter Technology.

Chavez Juan D JD Keller Andrew A Wippel Helisa H HH Mohr Jared P JP Bruce James E JE

Analytical chemistry 20211209 50

Chemical cross-linking with mass spectrometry (XL-MS) has emerged as a useful technique for interrogating protein structures and interactions. When combined with quantitative proteomics strategies, protein conformational and interaction dynamics can be probed. Quantitative XL-MS has been demonstrated with the use of stable isotopes incorporated metabolically or into the cross-linker molecules. Isotope-labeled cross-linkers have primarily utilized deuterium and rely on MS1-based quantitation of p ...[more]

PMID: 34882395

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quantitative crosslinking of ADP treated mitochondria isolated from HEK293 cells

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Project description:In vivo application of an extended set of six iqPIR reagents (6-plex iqPIR) allowing multiplexed quantitative interactome analysis of up to six biological samples in a single LC-MS acquisition. Multiplexed iqPIR is demonstrated on MCF-7 breast cancer cells treated with five different Hsp90 inhibitors revealing large scale protein conformational and interaction changes specific to the molecular class of the inhibitors.

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Mechanism of pharmacochaperoning in ATP-sensitive potassium channels revealed by cryo-EM

Project description:ATP-sensitive potassium (K-ATP) channels composed of a pore-forming Kir6.2 potassium channel and a regulatory ABC transporter sulfonylurea receptor 1 (SUR1) regulate insulin secretion in pancreatic beta-cells to maintain glucose homeostasis. Mutations that impair channel folding or assembly prevent cell surface expression and cause congenital hyperinsulinism. Structurally diverse K-ATP inhibitors have been shown to act as pharmacochaperones to correct mutant channel expression, but the mechanism is unknown. Here, we compare cryoEM structures of K-ATP channels bound to pharmacochaperones glibenclamide, repaglinide, and carbamazepine. DSP cross-linking mass spectrometry was used to partially confirm cryoEM structures.

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Deciphering the architecture and interactome of the spliceosomal C complex using in vivo chemical crosslinking

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2022-10-13 | PXD023577 | Pride

Age induced interactome remodeling in muscle mitochondria

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2024-05-23 | PXD031643 | Pride

Mechanism of pharmacochaperoning in ATP-sensitive potassium channels revealed by cryo-EM

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Structural basis of human full-length kindlin-3 homotrimer in an auto-inhibited state

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2020-06-08 | PXD019110 | Pride

Dynamics of different mGBP as revealed by computational and experimental methods

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