With proteins, form is function
To understand protein function and mechanism of action, it is essential to determine protein complex assembly and structure.
Thermo Fisher Scientific leads the way in accelerating protein structure-function studies with its Integrative Structural Biology solutions, which are complementary mass spectrometry (MS) and cryo-EM techniques that characterize complex and dynamic structure-function relationships.
Solving the structure of large dynamic complexes requires integrating several complementary techniques, such as MS and cryo-EM density maps, in an approach that is known as integrative structural biology.
One such example takes advantage of structural proteomics MS tools to study the stoichiometry of KaiA, KaiB, and KaiC (components of the cyanobacterial circadian clock), monitoring these well-defined assemblies followed by their structural characterization using single-particle cryo-EM.1
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Biomolecular MS has significantly advanced and impacted the field of structural biology. At the intact protein level, native MS enables the study of protein assemblies in their native state through the analysis of non-covalent protein-protein and protein-ligand complexes. At the peptide level, LC-MS/MS analysis of protein proteolytic digests helps determine the amino acid sequence of proteins, allowing their subunits to be identified from a proteome database.
Thermo Scientific Orbitrap MS solutions enable both peptide and protein-centric strategies that deliver insights into a multitude of biochemical and structural properties. Armed with superior quality data from proven HRAM Orbitrap systems, you can confidently analyze samples with increasing analytical depth, and deliver information that accelerates the journey from structure to function.
Applying mass spectrometry to study structure-function
Technology developments in MS have given rise to several applications of structural biology, both at the single protein and protein complex level. The primary advantages of MS-based techniques are the ability to perform experiments at proteome scale, to analyze proteins in their native biological state, and to reduce the minimum required sample size.
Protein structure analysis with mass spectrometry categories
Determining the shape and domain structure of a protein can provide evidence for enzyme mechanism and predict the function for uncharacterized proteins. The study of 3-D structure is also the starting point for further studies in protein-ligand interactions, posttranslational modifications, and evidence for transmembrane domains.
Protein-ligand interactions are involved in a plethora of biological functions, from protein transcription to translation and signal transduction. Information on binding kinetics, interaction relationships, and binding forces is typically uncovered during protein-ligand interaction experiments.
The majority of proteins undergo some level of posttranslational modification of their amino acid residues. These PTMs regulate interactions between proteins, nucleic acids, lipids, and other cofactors and can occur at any moment of the "life cycle" of a protein, influencing its function during biological processes such as catalysis, protein–protein interaction, and degradation.
An essential part of any proteomics workflow is identification of proteins. Such identification might include a focus on just one protein, or the profiling of hundreds of protein components in a single biological sample.
Many cellular processes are regulated by protein complexes. By identifying the stoichiometry of the individual components within the protein complex, one gains an understanding of that molecule's overall function.
While many molecules perform their functions independently, the vast majority of proteins interact with one another during biological activity. These interactions control cell processes, including protein modification, transport, folding, signaling, and cell cycling.
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The Thermo Scientific Orbitrap Fusion Lumos Tribrid Mass Spectrometer is the most advanced mass spectrometry system that can be upgraded to even higher resolution (1M), additional dissociation capabilities (UVPD and ETD), and a unique internal calibration (IC).
In collaboration with the editors of Science Magazine, we invite you to download this comprehensive booklet, containing articles from the Science family of journals, perspectives on integrative structural biology, and interviews with leading researchers.
We describe the preparation of stoichiometrically well-defined assemblies of KaiCB and KaiCBA, as monitored by native mass spectrometry, allowing for a structural characterization by single-particle cryo-electron microscopy and mass spectrometry.
- Snijder, J., Schuller, J.M., Wiegard, A., Lössl, P., Schmelling, N., Axmann, I.M., Plitzko, J.M., Förster, F. and Heck, A.J., 2017.
Structures of the cyanobacterial circadian oscillator frozen in a fully assembled state.
Science, 355(6330), pp.1181-1184.