The mechanism and energetics of the dynein priming stroke.

Dyneins are an AAA+ motor responsible for motility and force generation toward the minus end of microtubules. Dynein motility is powered by nucleotide-dependent transitions of its linker domain, which transitions between straight (post-powerstroke) and bent (pre-powerstroke) conformations. To understand the dynamics and energetics of the linker, we performed all-atom molecular dynamics simulations of human dynein-2 primed for its power stroke. Simulations revealed that the linker can adopt either a bent conformation or a semi-bent conformation, separated by a 5.7 kT energy barrier. The linker cannot switch back to its straight conformation in the pre-powerstroke state due to a steric clash with the AAA+ ring. Simulations also showed that an isolated linker has a free energy minimum near the semi-bent conformation in the absence of the AAA+ ring, indicating that the linker stores energy as it bends and releases this energy during the powerstroke.

SARS-CoV-2 Delta Variant Decreases Nanobody Binding and ACE2 Blocking Effectivity.

The Delta variant spreads more rapidly than previous variants of SARS-CoV-2. This variant comprises several mutations on the receptor-binding domain (RBDDelta) of its spike glycoprotein, which binds to the peptidase domain (PD) of angiotensin-converting enzyme 2 (ACE2) receptors in host cells. The RBD-PD interaction has been targeted by antibodies and nanobodies to prevent viral infection, but their effectiveness against the Delta variant remains unclear. Here, we investigated RBDDelta-PD interactions in the presence and absence of nanobodies H11-H4, H11-D4, and Ty1 by performing 21.8 µs of all-atom molecular dynamics simulations. Unbiased simulations revealed that Delta variant mutations strengthen RBD binding to ACE2 by increasing the hydrophobic interactions and salt bridge formation, but weaken interactions with H11-H4, H11-D4, and Ty1. Among these nanobodies H11-H4 and H11-D4 bind RBD without overlapping ACE2. They were unable to dislocate ACE2 from RBDDelta when bound side by side with ACE2 on RBD. Steered molecular dynamics simulations at comparable loading rates to high-speed atomic force microscopy (AFM) experiments estimated lower rupture forces of the nanobodies from RBDDelta compared to ACE2. Our results suggest that existing nanobodies are less effective to inhibit RBDDelta-PD interactions and a new generation of nanobodies is needed to neutralize the Delta variant.

Conformational transition of SARS-CoV-2 spike glycoprotein between its closed and open states.

In 2020, the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected millions of people worldwide and caused the coronavirus disease 2019 (COVID-19). Spike (S) glycoproteins on the viral membrane bind to ACE2 receptors on the host cell membrane and initiate fusion, and S protein is currently among the primary drug target to inhibit viral entry. The S protein can be in a receptor inaccessible (closed) or accessible (open) state based on down and up positions of its receptor-binding domain (RBD), respectively. However, conformational dynamics and the transition pathway between closed to open states remain unexplored. Here, we performed all-atom molecular dynamics (MD) simulations starting from closed and open states of the S protein trimer in the presence of explicit water and ions. MD simulations showed that RBD forms a higher number of interdomain interactions and exhibits lower mobility in its down position than its up position. MD simulations starting from intermediate conformations between the open and closed states indicated that RBD switches to the up position through a semi-open intermediate that potentially reduces the free energy barrier between the closed and open states. Free energy landscapes were constructed, and a minimum energy pathway connecting the closed and open states was proposed. Because RBD-ACE2 binding is compatible with the semi-open state, but not with the closed state of the S protein, we propose that the formation of the intermediate state is a prerequisite for the host cell recognition.

Thermodynamic first law efficiency of membrane proteins.

Proteins are nature's biomolecular machines. Proteins, such as transporters, pumps and motors, have complex function/operating-machinery/mechanisms, comparable to the macro-scaled machines that we encounter in our daily life. These proteins, as it is for their macro-scaled counterparts, convert (part of) other/various forms of energy into work. In this study, we are performing the first law analysis on a set of proteins, including the dopamine transporter, glycine transporters I and II, glutamate transporter, sodium-potassium pump and Ca2+ ATPase. Each of these proteins operates on a thermodynamic/mechanic cycle to perform their function. In each of these cycles, they receive energy from a source, convert part of this energy into work and reject the remaining part of the energy to the environment. Conservation of energy principle was applied to the thermodynamic/mechanic cycle of each protein, and thermodynamic first law efficiency was evaluated for each cycle, which shows how much of the energy input per cycle was converted into useful work. Interestingly, calculations based on experimental data indicate that proteins can operate under a range of efficiencies, which vary based on the extracellular and intracellular ion and substrate concentrations. The lowest observed first law efficiency was 50%, which is a very high value if compared to the efficiency of the macro-scaled heat engines we encounter in our daily lives.Communicated by Ramaswamy H. Sarma.