YouTube video by fab70smusic

AbstractWe report the first demonstration of ultraviolet photodissociation (UVPD) mass spectrometry achieved with light-emitting diodes (LEDs) externally interfaced to a linear ion-trap mass spectrometer. LEDs provide a low-cost, compact photon source that is technologically simpler to operate than the high-powered pulsed laser systems that are currently used in UVPD instruments. Mounting the LEDs external to the mass spectrometer provides improved flexibility for interchanging LEDs with different wavelengths. Here, we demonstrate 365 and 280 nm LED-UVPD on flavin mononucleotide, tryptophan, and the pentapeptide Leu-Enk. The UVPD fragments observed are in full agreement with fragments observed using pulsed lasers as the photon source.

YouTube video by blink-182 - Topic

We built and characterised a mass spectrometer capable of performing CID (both beam type and resonant type), UVPD, EID and ECD in an automated fashion during an LCMS type experiment. We exploited this...

A focused laser is used to make infrared multiphoton photodissociation (IRMPD) more efficient in a quadrupole ion trap mass spectrometer. Efficient (up to 100%) dissociation at the standard operating ...

Ultraviolet photodissociation (UVPD) has emerged as a powerful alternative to conventional collision-induced dissociation (CID) for peptide and protein sequencing in mass spectrometry-based proteomics. However, the UVPD efficiency depends on the presence of UV-absorbing chromophores within analytes. Here, we systematically investigate how N-terminal modification of peptides with aromatic chromophores influences the 193 nm UVPD efficiency and fragmentation behavior. Using NHS ester chemistry, human cell lysate digests were derivatized with a range of UV-absorbing aromatic labels and analyzed on an Omnitrap–Orbitrap platform equipped with a 193 nm ArF excimer laser. Compared with unmodified controls, derivatized peptides displayed a modestly increased fragmentation efficiency and a greater diversity of sequence-informative fragments, particularly b-type ions. Peptides lacking aromatic amino acids benefited most from N-terminal aromatic labeling, exhibiting enhanced fragmentation and sequence coverage. These findings demonstrate that incorporation of aromatic chromophores at the N-terminus can enhance UVPD performance and improve spectral interpretability, thereby expanding the applicability of UVPD in bottom-up proteomics workflows.

YouTube video by blink-182 - Topic

YouTube video by blink-182 - Topic

Journal of the American Society for Mass SpectrometryDOI: 10.1021/jasms.5c00130

YouTube video by blink-182 - Topic

YouTube video by blink-182 - Topic

The results of our recent development and characterization of UVPD, EID and ECD on the Omnitrap platform36 suggested that it could be deployed in an LC-MS configuration for the analysis of complex peptide mixtures. Given that the conditions in direct-infusion experiments from our earlier work, such as number of available ions, injection times and ion transfer logistics, are typically more relaxed than in automated LC-MS analysis, an investigation is required to determine the optimal parameters for all dissociation techniques. Direct-infusion experiments reported previously36 were focused on higher resolution and signal-to-noise ratio with no regard to duty cycle. Given that acquisition of spectra with this configuration has limited parallelization potential (Extended Data Fig. 1a), we initially concentrated on reducing scan length to increase speed of spectra acquisition to handle the complexity of proteomes. The Omnitrap design requires ions to be cooled through a gas pulse prior to any ion manipulation. The original design used a single gas valve that had a maximum repetition rate of 10 Hz (Extended Data Fig. 1b). To improve the maximum rate of the Omnitrap we implemented the use of two valves, operating alternately, for gas injection, which can potentially double the speed (Extended Data Fig. 1b)...

Directly probing the heterogeneous conformations of intracellular proteins within their native cellular environment remains a significant challenge in mass spectrometry (MS). Here, we establish an in-cell MS and ultraviolet photodissociation (UVPD) strategy that directly ejects proteins from living cells into a mass spectrometer, followed by 193 nm UVPD for structural analysis. Applying this approach to calmodulin (CaM), we reveal that it adopts more extended conformations within living cells compared with purified samples in vitro, highlighting the unique influence of intracellular environments on protein folding. Furthermore, UVPD analysis of calcium ion (Ca2+)-binding variants of CaM unveils not only the conformational heterogeneity induced by multiple Ca2+ modulations but also reveals distinct preferences of Ca2+ binding sites across different conformations. This strategy provides a powerful tool for interrogating the structure–function relationships of intracellular protein variants with sophisticated metal ion binding, paving the way for a deeper understanding of protein conformations within their native cellular context.

Proteins in cells are highly flexible and often exist in multiple conformations, each with unique abilities to bind ligands.

To date, collision-induced dissociation and methods based on electron transfer dissociation are considered the ‘best’ approaches for the mass spectrometry analysis of N- and O-glycopeptides, respectiv...

YouTube video by blink-182 - Topic

To date, collision-induced dissociation and methods based on electron transfer dissociation are considered the standard approaches for the mass spectrometry analysis of N- and O-glycopeptides, respectively, allowing for identification of both peptide and glycan compositions. In recent years, alternative fragmentation methods such as ultraviolet photodissociation (UVPD) and more energetic versions of electron-based techniques (such as electron ionisation dissociation, EID) have been shown to be useful for the analysis of glycopeptides, producing rich information on the glycopeptide structure, including types of glycosidic linkages. We evaluated ultraviolet photodissociation (UVPD), electron ionization dissociation (EID), electron capture dissociation (ECD), and activated-ion ECD (AI-ECD) using an Orbitrap-Omnitrap hybrid for LCMS analysis of complex N-glycopeptides. Both UVPD and EID generated extensive peptide, glycosidic, and cross-ring fragments, enabling detailed structural characterization. While ECD alone produced few glycopeptide identifications, AI-ECD significantly improved yields through supplemental vibrational activation. UVPD and EID achieved comparable identification efficiencies to stepped collisional dissociation and provided additional linkage information. These results establish the Omnitrap as a powerful platform for comprehensive glycoproteomic analysis and highlight the need for enhanced computational tools to interpret complex UVPD and EID spectra.