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Protein dimerization is one of the most widespread and functionally important forms of protein–protein interaction. Proteins rarely act in isolation in a biological environment; instead, they associate as homodimers, heterodimers or higher-order oligomers to form functional assemblies. This self-assembly occurs in every cellular compartment, from membranes to the nucleus and cytosol, and regulates processes ranging from enzymatic activation and signal transduction to pathogenic pathways. A dimer observed in a crystal structure, however, may represent either a genuine biological assembly or a contact created by crystal packing. A crystal structure provides detailed information about molecular architecture, but it does not by itself establish whether two molecules associate under relevant solution conditions, how strong that interaction is, or whether the subunits retain the same arrangement outside the lattice. Distinguishing between these possibilities is therefore an important task in structural biology, biophysics and pharmaceutical research, where protein–protein interactions can influence function, stability and the binding of potential drug candidates.

Protein dimerization occurs when two protein molecules associate to form a dimer. A crystal structure can reveal how two protein molecules fit together, identifying an interface that may influence protein function or drug discovery. However, a dimer observed in a crystal does not necessarily represent the dominant assembly in solution. Crystal packing can stabilize contacts that are weak, transient, or absent under physiologically relevant conditions. For pharmaceutical researchers, this distinction matters because a proposed dimer interface may form a binding site, regulate protein activity, or provide a target for small molecules. Before using the structure to guide drug design, researchers need to establish whether the protein associates in solution and whether the arrangement of its subunits is consistent with the proposed model.

Pulsed EPR spectroscopy is a form of electron paramagnetic resonance spectroscopy that excites unpaired electrons with short, strong microwave pulses and records how they respond over time, instead of sweeping a weak, constant microwave field. This article is for protein scientists, structural biologists and drug discovery teams. It explains what pulsed EPR measures, how it differs from continuous-wave EPR, which experiments it can run, and how pulsed dipolar spectroscopy turns two spin labels into a distance distribution.

Few tools have changed structural biology as quickly as AlphaFold2. Within a few years of its release, AlphaFold2 had become an important starting point for structural studies of many proteins. However, transporters, receptors, channels and many enzymes function through conformational changes, and ligand binding can shift proteins between different structural states. A single predicted model, however confident, represents one structural state rather than the full range of conformations a protein may populate. This is where Electron Paramagnetic Resonance (EPR) spectroscopy comes in.

Allosteric regulation is a fundamental mechanism of cellular control, enabling proteins to respond to signals at one site and translate them into changes in activity at another, often distant, site. Proteins regulated in this way, known as allosteric proteins, are involved in a wide range of biological processes, including enzyme regulation, signal transduction, cell adhesion and transcription.

Electron paramagnetic resonance spectroscopy is a powerful way to study materials, molecules, and systems with unpaired electrons. It is often called EPR, ESR, electron spin resonance spectroscopy, or spin resonance spectroscopy.

A protein conformational change is often described as a transition from one structural state to another: open to closed, inactive to active, or apo to ligand-bound. That language is useful for describing structural differences, but it can obscure the physical problem that determines how a protein behaves in solution.

Artificial intelligence has changed the starting point for structural biology. Protein structure prediction tools such as AlphaFold can generate highly accurate structural models for many proteins, including proteins for which experimentally determined structures are unavailable.

Structure-based drug design uses the three-dimensional structure of a target protein to identify functional binding sites and design drug molecules that complement their shape, size, and electrostatic properties. This approach has enabled major advances in medicine, including HIV protease inhibitors, antibiotics, and targeted cancer therapies such as imatinib.

A high-resolution cryo-EM map can still leave important regions unresolved. Here's what missing density may and may not tell you about protein structure, flexibility, and conformational heterogeneity.

Electron paramagnetic resonance (EPR) spectroscopy is one of the most effective methods for studying membrane protein structure and dynamics. Unlike X-ray crystallography and cryo-electron microscopy, which provide static structural snapshots, EPR measures conformational ensembles and molecular motions under native-like conditions. This makes it particularly valuable for investigating GPCRs, ion channels, and membrane transporters involved in drug discovery.

The Rocky Mountain Conference on Magnetic Resonance remains one of the premier gatherings for the EPR community. This year, High Q Technologies will share new advances in automated EPR spectroscopy with the FATHOM system, highlighting improved sensitivity, phase stability, and applications in structural biology and challenging membrane protein research.