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Rise to the structural biology challenges of today and tomorrow
Biological insights often require the study of systems as they interact. Structural biology aims to study molecules as close to in vivo as possible to accomplish this goal. The Thermo Scientific Orbitrap Tribrid Apex Structural Biology Mass Spectrometer is our most flexible instrument, making it ideal for problems that researchers in this field will face. It excels in experiments ranging from native protein characterization, crosslinking, hydrogen-deuterium exchange mass spectrometry (HDX-MS), and ligand binding, while maintaining flexibility and ease-of-use.
Here we introduce the first commercial IR-based laser, for more efficient fragmentation using IR-ETD, and better desolvation for native proteins. Expand your capabilities with complex native samples with Direct Mass Technology mode and combine it with all the versatility the Orbitrap Tribrid MS platform offers. Delivering yet another innovative solution for real experimental problems. Let Thermo Fisher Scientific be your partner now, and your future endeavors.
What scientists are saying about Orbitrap Tribrid Apex Structural Biology MS
"I am very excited about the Orbitrap Tribrid Apex Structural Biology MS. Its single-ion detection capability significantly expands the dynamic range of detecting protein complexes, enabling detection of low-abundance but biologically critical signaling complexes.
Combined with multistage MSn fragmentation in both ensemble and single-ion modes, this platform positions native mass spectrometry as a powerful discovery tool for discerning how key membrane-associated signaling complexes drive cell biology of human health and disease."
Kallol Gutpa, Ph.D., Associate Professor, Yale University
"The array of MS/MS techniques, now including an IR-laser, available on the Orbitrap Tribrid Apex Structural Biology mass spectrometer greatly expands characterization of large assemblies, especially ribonucleoproteins. The IR-laser enhances desolvation for greater accuracy in mass determination of the assemblies. Additionally, IRMPD in combination with ETD significantly boosts top-down and native top-down characterization of the subunits and intact assemblies."
Jared Shaw, Ph.D., Assistant Professor, University of Nebraska-Lincoln
Applications
Native MS: Maintain native structure through desolvation
The new IR-laser can be used to gently and efficiently assist in the desolation process for native complexes. Below we see Aquaporin Z tetramer, without any activation, the MS1 has the characteristic tailing of solvent/adduct adhesion. As we see with a 10% IR-laser pulse, there is a 10X increase in signal with no tailing nor charge shift, implying native structure is maintained, all while improving signal by shifting adduct and solvent peaks to only the true mass.
Advanced antibody Fab sequencing
Successfully characterize mAbs including their full proteoforms and nearly all their variable regions after IdeS digestion. Direct Mass Technology mode assists in increasing sequence coverage by directly gaining mass domain information from complex top-/middle-down spectral information. This translates to more sequence coverage especially in the middle of the protein’s sequence where it can often be difficult for traditional ensemble-based methods to deconvolute.
IdeS digestion followed by reduction generates Fd and light chain fragments from intact IgG, enabling high-confidence mass spectrometry sequencing of antibody variable regions using Direct Mass Technology mode.
Native MS complex characterization
The IR-laser maintains native structure and improves radical fragmentation via ETD. The dense fragment spectrum can be simplified for matching by deconvolution by utilizing PTCR. Here we show the fragmentation pattern from native Malate Dehydrogenase (MDH) in its dimer form. Utilizing IR-assisted EThcD, we see 60% sequence coverage from MS2 scans, but can increase that to 75% when utilizing MS3 and PTCR to spread out the fragment ions for easier deconvolution. Both methods allow for deeper context into native proteoforms.
HDX-MS: Improved resolution for single-residue structural insights
Single-residue hydrogen-deuterium exchange mass spectrometry (HDX-MS) provides uniquely detailed insight into protein structure, dynamics, and interactions by localizing exchange behavior at the amino-acid level, enabling resolution of subtle conformational changes inaccessible to peptide-level HDX-MS. Achieving this capability requires fragmentation methods that preserve solution-phase deuterium labeling, as gas-phase hydrogen scrambling can obscure residue-specific information.
The Orbitrap Tribrid Apex Structural Biology MS builds on the soft ion optics of previous models and offers multiple fragmentation techniques suitable for residue-level HDX-MS including the new IR-laser option that allows for the lowest possible level of scrambling, enabling researchers to gain the highest resolution possible for structural information.
Scrambling evaluation using P1 model peptide (shown on left). IR-assisted ETD and EThcD reduce scrambling compared with HCD, helping preserve deuterium labeling for more confident residue-level HDX-MS analysis.
Improved quantifiable crosslinking-MS capabilities
Quantification alongside crosslinking data is essential in structural biology because it reveals not only which interactions occur, but how strongly and how frequently they occur under specific biological conditions. By measuring relative crosslink abundances, researchers can distinguish stable structural features from dynamic or transient interactions, enabling more accurate modeling of conformational changes, interaction networks, and functional states of protein complexes. The improved parallelization on the Orbitrap Tribrid Apex Structural Biology MS increases ion injection times, increasing data quality. This paired with the new IR-laser for greater Tandem Mass Tags (TMT) reporter ion signal to noise results in a 15% improvement in quantifiable crosslinks (XLs). Giving scientists more insights into their biological questions.
FAQs
In relation to structural biology applications, the IR-laser can improve many mass spectrometry-based experiments. First, the laser can be used to gently desolvate complexes while maintaining their native structure. Next, the IR-laser can be coupled with radical fragmentation from ETD (IR-ETD/EThcD) which shows a large improvement in the signal-to-noise of c-/z- fragment ions. Finally, when using quantitation with TMT, the IR-laser improves signal-to-noise for TMT reporter ions, enabling more accurate quantitation.
Direct Mass Technology mode is a method to take parallel individual ion measurements in the Orbitrap and get accurate mass determination outside of the m/z domain. This is a very useful technique for any application in which the analyte is not able to be resolved via traditional ensemble methods. For example, protein complexes, biotherapeutics, or top-down fragmentation data. Simply toggle on the mode, collect the raw data, and upload into STORIboard software. Learn more about Direct Mass Technology.
Each protein and protein complex is unique and can require individual optimization for best results. However, we emphasize useability on this platform, and so premade template methods are provided to allow for appropriate starting points on all major applications within structural biology.
For Research Use Only. Not for use in diagnostic procedures.