Imaging Large Multiprotein Complexes Using Native Nano DESI Mass Spectrometry
- Photo: Thermo Fisher Scientific: Imaging Large Multiprotein Complexes Using Native Nano DESI Mass Spectrometry
- Video: Chromatography & Mass Spectrometry Solutions: Introducing the Thermo Scientific Orbitrap Ascend Tribrid MS
Prof. Helen Cooper, Ph.D., an expert analytical chemist from the University of Birmingham, recently discussed the latest advancements in imaging large multiprotein complexes using native Nano-spray desorption electrospray ionization (nano-DESI) mass spectrometry during a webinar View on-demand webinar.
Background on native ambient mass spectrometry imaging
Native ambient mass spectrometry imaging allows scientists to visualize the spatial distribution of proteins and their complexes in thin tissue sections. This technology provides valuable insights into protein therapeutics, such as monoclonal antibodies. However, analyzing large multiprotein complexes using existing imaging technologies remains challenging.
Dr. Cooper discussed improvements in mass spectrometry imaging that enable the characterization of precise proteins present in multiprotein complexes. During this webinar, you will learn about the Cooper Lab’s experience using the Thermo Scientific Orbitrap Eclipse tribrid mass spectrometer and the Thermo Scientific Orbitrap Ascend structural biology tribrid mass spectrometer.
Key topics covered
- Orbitrap Ascend structural biology tribrid mass spectrometer: Learn how this instrument can accelerate native workflows with its capabilities for isolating ions up to m/z 8,000 in the quadrupole and detecting up to m/z 16,000 in the Orbitrap mass analyzer.
- Deriving cellular information: Understand how native mass spectrometry of protein complexes can provide cellular information.
- Combining imaging MS with native structural analysis: Discover how this combination can be used to provide proteoform-level information.
Presentation overview
The presentation begins by explaining the goal of combining native mass spectrometry and mass spectrometry imaging to obtain simultaneous information about spatial distribution and protein structure directly from tissue. This approach integrates three strands of mass spectrometry:
- Native mass spectrometry: Proteins are electrosprayed from non-denaturing conditions, retaining non-covalent interactions in the gas phase, keeping protein assemblies intact.
- Top-down mass spectrometry: Protein ions are fragmented to provide sequence information, identifying the protein and post-translational modifications.
- Ambient mass spectrometry imaging: Allows spatial analysis of proteins in thin tissue sections. Performing native nano DESI imaging on tissue sections involves using a liquid junction sampling technique. This method allows them to sample tissues without disrupting protein structures.
Case study: Metal-deficient SOD1 dimer in ALS
A significant case study involved imaging the metal-deficient SOD1 dimer in spinal cord tissue from a mouse model of amyotrophic lateral sclerosis (ALS). The study revealed an abundance of metal-deficient SOD1 dimer in regions of pathology, which correlates with ALS pathology. This was achieved using the Orbitrap Eclipse Tribrid Mass Spectrometer.
Instrumentation and achievements
Practical molecular weight limits
The Orbitrap Eclipse Tribrid Mass Spectrometer has been instrumental in achieving significant milestones:
- Identification of the largest protein assembly from tissue at 145 kilodaltons.
- Imaging of a 113 kilodalton aquaporin 0 tetramer.
Proton transfer charge reduction (PTCR)
PTCR has been utilized to determine intact masses and improve sensitivity. This technique allows the separation and identification of protein species from complex spectra, addressing the challenges of working with low-abundance signals and complex mass spectra.
Advancements with Orbitrap Ascend structural biology MS
Data independent acquisition (DIA)-PTCR
The DIA- PTCR method involves scanning along the m/z range and subjecting slices to PTCR. This method has enabled the detection of approximately 100 proteoforms, providing comprehensive proteoform discovery.
Imaging higher molecular weight proteins
The Orbitrap Ascend structural biology tribrid MS led to the imaging of higher molecular weight proteins, such as the 231 kilodalton pyruvate kinase tetramer. This represents a significant improvement, more than doubling the previous molecular weight range for mass spectrometry imaging of proteins.
Conclusion
Native ambient mass spectrometry imaging is a comprehensive platform for in situ native protein characterization and imaging. The advancements provided by the Orbitrap Ascend structural biology tribrid mass spectrometer have enabled significant progress in this field, doubling the molecular weight range for imaging and facilitating the characterization of larger macromolecular machines.
This progress highlights the potential for deeper insights into protein structures and their functions within biological tissues, paving the way for advancements in understanding complex biological systems and developing new therapeutic strategies.
Learn more at www.thermofisher.com/OrbitrapAscendStructuralBiology
Visit us on LinkedIn: #StructuralBiology #NativeMS #nanoDESI #OrbitrapAscendStructuralBiology
_s.webp)
