Dataset Information

"In-gel cross-linking mass spectrometry (IGX-MS) of proteins and protein complexes"

ABSTRACT: Cross-linking mass spectrometry has developed into an important method to study protein structures and interactions. The in-solution cross-linking workflows involve time and sample consuming steps and do not provide sensible solutions for differentiating cross-links obtained from co-occurring protein oligomers, complexes, or conformers. Here we developed a cross-linking workflow combining blue native PAGE with in-gel cross-linking mass spectrometry (IGX-MS). This workflow circumvents steps, such as buffer exchange and cross-linker concentration optimization. Additionally, IGX-MS enables the parallel analysis of co-occurring protein complexes using only small amounts of sample. Another benefit of IGX-MS observed by experiments on GroEL and purified bovine heart mitochondria, is the substantial reduction of artificial over-length cross-links when compared to in-solution cross-linking. We next used IGX-MS to investigate the complement components C5, C6, and their hetero-dimeric C5b6 complex. The obtained cross-links were used to generate a refined structural model of the complement component C6, resembling C6 in its inactivated state. This finding shows that IGX-MS can be used to provide new insights into the initial stages of the terminal complement pathway.

INSTRUMENT(S): Orbitrap Fusion Lumos, Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human) Bos Taurus (bovine) Escherichia Coli

TISSUE(S): Heart, Cell Culture, Blood

SUBMITTER: Johannes Hevler

LAB HEAD: Albert J. R Heck

PROVIDER: PXD020014 | Pride | 2020-12-14

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	20191219_L1_Ag1_Lukas006_SA_GroEL_DSSO_100nM_1-2_Crosslinks.txt	Txt
	20191219_L1_Ag1_Lukas006_SA_GroEL_DSSO_100nM_1.raw	Raw
	20191219_L1_Ag1_Lukas006_SA_GroEL_DSSO_100nM_2-2_Crosslinks.txt	Txt
	20191219_L1_Ag1_Lukas006_SA_GroEL_DSSO_100nM_2.raw	Raw
	20191219_L1_Ag1_Lukas006_SA_GroEL_DSSO_100nM_3-2_Crosslinks.txt	Txt

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"In-gel cross-linking mass spectrometry (IGX-MS) of proteins and protein complexes"

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