Viruses are highly ordered supramolecular complexes that have evolved to propagate by hijacking the host cell's machinery. Although viruses are very diverse, spreading through cells of all kingdoms of life, they share common functions and properties. Next to the general interest in virology, fundamental viral mechanisms are of growing importance in other disciplines such as biomedicine and (bio)nanotechnology. However, in order to optimally make use of viruses and virus‐like particles, for instance as vehicle for targeted drug delivery or as building blocks in electronics, it is essential to understand their basic chemical and physical properties and characteristics. In this context, the number of studies addressing the mechanisms governing viral properties and processes has recently grown drastically. This review summarizes a specific part of these scientific achievements, particularly addressing physical virology approaches aimed to understand the self‐assembly of viruses and the mechanical properties of viral particles. Using a physicochemical perspective, we have focused on fundamental studies providing an overview of the molecular basis governing these key aspects of viral systems. This article is categorized under: Biology‐Inspired Nanomaterials > Protein and Virus‐Based Structures Nanotechnology Approaches to Biology > Nanoscale Systems in Biology

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Single-molecule fluorescence optical tweezers studies combined with high-speed AFM reveal early events in the HBV life cycle.

Macrophages are well known for their role in immune responses and tissue homeostasis. They can polarize towards various phenotypes in response to biophysical and biochemical stimuli. However, little is known about the early kinetics of macrophage polarization in response to single biophysical or biochemical stimuli. Our approach, combining optical tweezers, confocal fluorescence microscopy, and microfluidics, allows us to isolate single macrophages and follow their immediate responses to a biochemical stimulus in real‐time. This strategy enables live‐cell imaging at high spatiotemporal resolution and omits surface adhesion and cell–cell contact as biophysical stimuli. The approach is validated by successfully following the early phase of an oxidative stress response of macrophages upon phorbol 12‐myristate 13‐acetate (PMA) stimulation, allowing detailed analysis of the initial macrophage response upon a single biochemical stimulus within seconds after its application, thereby eliminating delay times introduced by other techniques during the stimulation procedure. Hence, an unprecedented view of the early kinetics of macrophage polarization is provided.

The secondary active transporter CitS shuttles citrate across the cytoplasmic membrane of gram-negative bacteria by coupling substrate translocation to the transport of two Na+ ions. Static crystal structures suggest an elevator type of transport mechanism with two states: up and down. However, no dynamic measurements have been performed to substantiate this assumption. Here, we use high-speed atomic force microscopy for real-time visualization of the transport cycle at the level of single transporters. Unexpectedly, instead of a bimodal height distribution for the up and down states, the experiments reveal movements between three distinguishable states, with protrusions of ∼0.5 nm, ∼1.0 nm, and ∼1.6 nm above the membrane, respectively. Furthermore, the real-time measurements show that the individual protomers of the CitS dimer move up and down independently. A three-state elevator model of independently operating protomers resembles the mechanism proposed for the aspartate transporter GltPh. Since CitS and GltPh are structurally unrelated, we conclude that the three-state elevators have evolved independently.