7 research outputs found
Effects of Lorentz invariance violation on cosmic ray photon emission and gamma ray decay processes
In this work, we use Lorentz invariance violation (LIV) introduced as a
generic modification to particle dispersion relations to study some
consequences of single photon emission, known as vacuum Cherenkov radiation,
and photon decay processes in cosmic and gamma rays. These processes are
forbidden in a Lorentz invariant theory but allowed under the hypothesis of
LIV. We show that the emission rate have a dependency on the cosmic ray primary
mass and the electric charge that could modify the UHECR spectrum. Furthermore,
LIV dramatically enhances photon decay into an electro-positron pair above
certain energy threshold. This last effect can then be used to set limits to
the LIV energy scale from the direct observation of very high energy cosmic
photon events by telescopes of gamma-rays.Comment: Proceedings of the 35th International Cosmic Ray Conference (ICRC
2017), Busan, Kore
Structural Analysis of an Intact Monoclonal Antibody by Online Electrochemical Reduction of Disulfide Bonds and Fourier Transform Ion Cyclotron Resonance Mass Spectrometry
Structural confirmation and quality
control of recombinant monoclonal
antibodies (mAbs) by top-down mass spectrometry is still challenging
due to the size of the proteins, disulfide content, and post-translational
modifications such as glycosylation. In this study we have applied
electrochemistry (EC) to overcome disulfide bridge complexity in top-down
analysis of mAbs. To this end, an electrochemical cell was coupled
directly to an electrospray ionization (ESI) source and a Fourier
transform ion cyclotron resonance (FTICR) mass spectrometer (MS) equipped
with a 15 T magnet. By performing online EC-assisted reduction of
interchain disulfide bonds in an intact mAb, the released light chains
could be selected for tandem mass spectrometry (MS/MS) analysis without
interference from heavy-chain fragments. Moreover, the acquisition
of full MS scans under denaturing conditions allowed profiling of
all abundant mAb glycoforms. Ultrahigh-resolution FTICR-MS measurements
provided fully resolved isotopic distributions of intact mAb and enabled
the identification of the most abundant adducts and other interfering
species. Furthermore, it was found that reduction of interchain disulfide
bonds occurs in the ESI source dependent on capillary voltage and
solvent composition. This phenomenon was systematically evaluated
and compared with the results obtained from reduction in the electrochemical
cell
Top-Down MALDI-In-Source Decay-FTICR Mass Spectrometry of Isotopically Resolved Proteins
An
accurate mass measurement of a known protein provides information
on potential amino acid deletions and post-translational modifications.
Although this field is dominated by strategies based on electrospray
ionization, mass spectrometry (MS) methods using matrix-assisted laser
desorption/ionization (MALDI) have the advantage of yielding predominantly
singly charged precursor ions, thus avoiding peak overlap from different
charge states of multiple species. Such MALDI-MS methods require mass
measurement at ultrahigh resolution, which is provided by Fourier
transform ion cyclotron resonance (FTICR) mass analyzers. Recently,
using a MALDI-FTICR-MS platform equipped with a 15 T magnet, we reported
on the mass analysis of intact human serum peptides and small proteins
with isotopic resolution up to ∼15 kDa and identified new proteoforms
from an accurate measurement of mass distances. In the current study,
we have used this FTICR system after an upgrade with a novel dynamically
harmonized ICR cell, i.e., ParaCell, for mapping isotopically resolved
intact proteins up to about 17 kDa and performed top-down MALDI in-source
decay (ISD) analysis. Standard proteins myoglobin (<i>m</i>/<i>z</i>-value 16 950) and ribonuclease B (<i>m</i>/<i>z</i>-value 14 900) were measured
with resolving powers of 62 000 and 61 000, respectively.
Furthermore, it will be shown that (singly charged) MALDI-ISD fragment
ions can be measured at isotopic resolution up to <i>m</i>/<i>z</i>-value 12 000 (e.g., resolving power 39 000
at <i>m</i>/<i>z</i>-value 12 000) providing
more reliable identifications. Moreover, examples are presented of
pseudo-MS<sup>3</sup> experiments on ISD fragment ions from RNase
B by collisional-induced dissociation (CID)
Identification of New Apolipoprotein-CIII Glycoforms with Ultrahigh Resolution MALDI-FTICR Mass Spectrometry of Human Sera
Apolipoprotein-CIII
(apoCIII) is an abundant blood glycoprotein
associated with lipoprotein particles. Three different glycoforms
have been described, all containing a mucin-type core-1 <i>O</i>-glycosylation with either zero, one or two sialic acids. Changes
in the relative abundance of these glycoforms have been observed in
a variety of different pathologies. In this study, ultrahigh resolution
15T MALDI Fourier transform ion cyclotron resonance (FTICR) MS was
used to analyze apoCIII isoforms in serum protein profiles. For this
purpose, serum proteins were purified using both a fully automated
RPC18-based magnetic bead method and an RPC4 cartridge-based solid
phase extraction method. Six new apoCIII isoforms were identified
with low-ppm mass measurement errors and ultrahigh precision. These
were characterized by more complex glycan moieties that are fucosylated
instead of sialylated. To confirm the glycan moiety and localize the
glycosylation site, top-down ESI-FTICR-MS/MS and bottom-up LC-ion
trap MS/MS were used. A large variation in the presence and abundance
of the fucosylated isoforms was found in a set of 96 serum samples.
These findings of fucosylated apolipoprotein-CIII isoforms warrant
further research to elucidate the implications these glycoforms may
have for the plethora of studies where alterations in apoCIII have
been linked to the development of many different pathologies
Additional file 3: of IgA N- and O-glycosylation profiling reveals no association with the pregnancy-related improvement in rheumatoid arthritis
Supplementary figures. Figures S1 and S2 Values depicted in graphs from data in Additional file 2: Table S3. (DOCX 248 kb
Top-Down FTICR MS for the Identification of Fluorescent Labeling Efficiency and Specificity of the Cu-Protein Azurin
Fluorescent protein labeling has been an indispensable
tool in
many applications of biochemical, biophysical, and cell biological
research. Although detailed information about the labeling stoichiometry
and exact location of the label is often not necessary, for other
purposes, this information is crucial. We have studied the potential
of top-down electrospray ionization (ESI)-15T Fourier transform ion
cyclotron resonance (FTICR) mass spectrometry to study the degree
and positioning of fluorescent labeling. For this purpose, we have
labeled the Cu-protein azurin with the fluorescent label ATTO 655-N-hydroxysuccinimide(NHS)-ester
and fractionated the sample using anion exchange chromatography. Subsequently,
individual fractions were analyzed by ESI-15T FTICR to determine the
labeling stoichiometry, followed by top-down MS fragmentation, to
locate the position of the label. Results showed that, upon labeling
with ATTO 655-NHS, multiple different species of either singly or
doubly labeled azurin were formed. Top-down fragmentation of different
species, either with or without the copper, resulted in a sequence
coverage of approximately 50%. Different primary amine groups were
found to be (potential) labeling sites, and Lys-122 was identified
as the major labeling attachment site. In conclusion, we have demonstrated
that anion exchange chromatography in combination with ultrahigh resolution
15T ESI-FTICR top-down mass spectrometry is a valuable tool for measuring
fluorescent labeling efficiency and specificity
Typing <i>Pseudomonas aeruginosa</i> Isolates with Ultrahigh Resolution MALDI-FTICR Mass Spectrometry
The introduction of standardized
matrix-assisted laser desorption/ionization
time-of-flight mass spectrometry (MALDI-TOF MS) platforms in the medical
microbiological practice has revolutionized the way microbial species
identification is performed on a daily basis. To a large extent, this
is due to the ease of operation. Acquired spectra are compared to
profiles obtained from cultured colonies present in a reference spectra
database. It is fast and reliable, and costs are low compared to previous
diagnostic approaches. However, the low resolution and dynamic range
of the MALDI-TOF profiles have shown limited applicability for the
discrimination of different bacterial strains, as achieved with typing
based on genetic markers. This is pivotal in cases where certain strains
are associated with, e.g., virulence or antibiotic resistance. Ultrahigh
resolution MALDI-FTICR MS allows the measurement of small proteins
at isotopic resolution and can be used to analyze complex mixtures
with increased dynamic range and higher precision than MALDI-TOF MS,
while still generating results in a similar time frame. Here, we propose
to use ultrahigh resolution 15T MALDI-Fourier transform ion cyclotron
resonance (FTICR) MS to discriminate clinically relevant bacterial
strains after species identification performed by MALDI-TOF MS. We
used a collection of well characterized <i>Pseudomonas aeruginosa</i> strains, featuring distinct antibiotic resistance profiles, and
isolates obtained during hospital outbreaks. Following cluster analysis
based on amplification fragment length polymorphism (AFLP), these
strains were grouped into three different clusters. The same clusters
were obtained using protein profiles generated by MALDI-FTICR MS.
Subsequent intact protein analysis by electrospray ionization (ESI)-collision-induced
dissociation (CID)-FTICR MS was applied to identify protein isoforms
that contribute to the separation of the different clusters, illustrating
the additional advantage of this analytical platform