Source 1primary paper2020Journal of the American Society for Mass Spectrometry
Toolkit/top-down mass spectrometry
top-down mass spectrometry
Also known as: top-down MS
Taxonomy: Technique Branch / Method. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
Top-down mass spectrometry is an assay method for de novo characterization of covalent insulin dimers formed under Fe2+ incubation or UV light stress. In the cited workflow, combined EThcD and 213 nm UVPD enabled identification of cross-link chemical composition and residue-level cross-link sites.
Usefulness & Problems
Why this is useful
This method is useful for structural characterization of covalent protein dimers when both the cross-link chemistry and the residue-level linkage sites must be determined. In the cited insulin application, it resolved cross-link types and chain-to-chain cross-link sites using fragmentation-based top-down analysis.
Source:
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Source:
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Source:
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Problem solved
It addresses the problem of identifying the chemical nature and exact residue connectivity of covalent insulin dimers generated by Fe2+ incubation or UV light stress. The study specifically showed that MS3 analysis of informative MS2 fragments enabled residue-level site assignment for both dimer types.
Problem links
Need precise spatiotemporal control with light input
DerivedTop-down mass spectrometry is an assay method used for de novo characterization of covalent insulin dimers. In the cited workflow, it identified cross-link chemical types and residue-level cross-link sites in dimers formed by Fe2+ incubation or UV light stress using combined fragmentation strategies.
Taxonomy & Function
Primary hierarchy
Technique Branch
Method: A concrete measurement method used to characterize an engineered system.
Mechanisms
backbone fragmentationdisulfide bond cleavageHeterodimerizationPhotocleavagePhotocleavagePhotocleavageTarget processes
No target processes tagged yet.
Input: Light
Implementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationimplementation constraint: multi component delivery burdenimplementation constraint: spectral hardware requirementoperating role: sensorswitch architecture: cleavageswitch architecture: multi componentswitch architecture: recruitment
The reported workflow used top-down MS with combined EThcD and 213 nm UVPD fragmentation. MS3 analysis of selected MS2 fragments was required for residue-level site assignment in both dimers, although the Fe2+-induced dimer could be localized by UVPD-MS2 without MS3. The analytes were insulin dimers generated by Fe2+ incubation or UV light stress, and interchain disulfide bond cleavage contributed to localization in the MS3 workflow.
MS2 alone was not sufficient for cross-link site identification in the UV light-induced insulin dimer, so deeper fragmentation was required. The evidence is limited to covalent insulin dimers from one study, and broader performance across other proteins, cross-link classes, or sample types was not provided.
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
UVPD at MS2 provided cross-link site identification for the Fe2+-induced insulin dimer without MS3, but MS2 was not sufficient for cross-link site identification in the UV light-induced dimer.
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was implicated in the Fe2+-induced dimer cross-link, explaining why UVPD-MS2 effectively fragmented the cross-link and nearby backbone bonds.
The UV chromophoric side chain of Phe1 was indicated in the cross-link, explaining why UVPD-MS2 was effective in fragmenting the cross-link and nearby backbone bonds.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Combined EThcD and 213 nm UVPD facilitated identification of the chemical composition of cross-links in insulin dimers.
The combined utilization of EThcD and 213 nm ultraviolet photodissociation (UVPD) facilitated identification of the chemical composition of the cross-links.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
HCD, EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
Our results demonstrated that higher-energy collisional dissociation (HCD), EThcD, and UVPD combined with MS3 were powerful tools for direct de novo characterization of cross-linked insulin dimers.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
At the MS2 level, EThcD efficiently cleaved interchain disulfide bonds in insulin dimers to reveal cross-link connectivities between chains.
On the MS2 level, electron-transfer/higher-energy collision dissociation (EThcD) efficiently cleaved the interchain disulfide bonds in the dimers to reveal cross-link connectivities between chains.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
Approval Evidence
1 source4 linked approval claimsfirst-pass slug top-down-mass-spectrometry
de novo characterization in top-down mass spectrometry (MS) workflows
Source:
method capabilitysupports
Residue-level identification of cross-link sites between chains was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Identification of cross-link sites between chains at residue level was achievable for both dimers with MS3 analysis of MS2 fragments cleaved at the cross-link or additionally the interchain disulfide bonds.
Source:
method capabilitysupports
Top-down mass spectrometry workflows can identify cross-link types and sites in covalent insulin dimers induced by Fe2+ incubation or UV light stress.
we here demonstrate on two case studies of covalent insulin dimers, induced by Fe2+ incubation or ultraviolet (UV) light stress, that de novo characterization in top-down mass spectrometry (MS) workflows can identify cross-link types and sites
Source:
structural assignmentsupports
In the Fe2+-induced insulin dimer, Phe1 from both B-chains were cross-linked through a -CH2- linkage.
In the Fe2+-induced dimer, Phe1 from both B-chains were cross-linked through a -CH2-.
Source:
structural assignmentsupports
In the UV light-induced insulin dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain.
in the UV light-induced dimer, Tyr14 of the A-chain participated in an -O-S- cross-link in which the sulfur was derived either from Cys7 or Cys19 of the B-chain
Source:
Comparisons
Source-backed strengths
Combined EThcD and 213 nm UVPD facilitated identification of cross-link chemical composition in insulin dimers. Residue-level cross-link site identification was achieved for both dimers by MS3 analysis of MS2 fragments cleaved at the cross-link or additionally at interchain disulfide bonds. For the Fe2+-induced dimer, UVPD at MS2 alone was sufficient for cross-link site identification, indicating strong performance in at least one cross-link context.
Source:
UVPD provided identification of cross-link sites in the Fe2+-induced dimer without MS3, while cross-link site identification with MS2 was not possible for the UV light-induced dimer.
Compared with electron-electron double resonance spectroscopy
top-down mass spectrometry and electron-electron double resonance spectroscopy address a similar problem space.
Shared frame: same top-level item type; shared mechanisms: heterodimerization; same primary input modality: light
Compared with electron-transfer/higher-energy collision dissociation
top-down mass spectrometry and electron-transfer/higher-energy collision dissociation address a similar problem space.
Shared frame: same top-level item type; shared mechanisms: heterodimerization, photocleavage; same primary input modality: light
Strengths here: looks easier to implement in practice.
Compared with ultraviolet photodissociation
top-down mass spectrometry and ultraviolet photodissociation address a similar problem space.
Shared frame: same top-level item type; shared mechanisms: heterodimerization; same primary input modality: light
Ranked Citations
- 1.