The chloroplast protein HCF164 is predicted to be associated with Coffea SH9 resistance factor against Hemileia vastatrix.

To explore the connection between chloroplast and coffee resistance factors, designated as SH1 to SH9, whole genomic DNA of 42 coffee genotypes was sequenced, and entire chloroplast genomes were de novo assembled. The chloroplast phylogenetic haplotype network clustered individuals per species instead of SH factors. However, for the first time, it allowed the molecular validation of Coffea arabica as the maternal parent of the spontaneous hybrid "Híbrido de Timor". Individual reads were also aligned on the C. arabica reference genome to relate SH factors with chloroplast metabolism, and an in-silico analysis of selected nuclear-encoded chloroplast proteins (132 proteins) was performed. The nuclear-encoded thioredoxin-like membrane protein HCF164 enabled the discrimination of individuals with and without the SH9 factor, due to specific DNA variants linked to chromosome 7c (from C. canephora-derived sub-genome). The absence of both the thioredoxin domain and redox-active disulphide center in the HCF164 protein, observed in SH9 individuals, raises the possibility of potential implications on redox regulation. For the first time, the identification of specific DNA variants of chloroplast proteins allows discriminating individuals according to the SH profile. This study introduces an unexplored strategy for identifying protein/genes associated with SH factors and candidate targets of H. vastatrix effectors, thereby creating new perspectives for coffee breeding programs.

Allosteric communication in class A ß-lactamases occurs via cooperative coupling of loop dynamics.

Understanding allostery in enzymes and tools to identify it offer promising alternative strategies to inhibitor development. Through a combination of equilibrium and nonequilibrium molecular dynamics simulations, we identify allosteric effects and communication pathways in two prototypical class A ß-lactamases, TEM-1 and KPC-2, which are important determinants of antibiotic resistance. The nonequilibrium simulations reveal pathways of communication operating over distances of 30 Å or more. Propagation of the signal occurs through cooperative coupling of loop dynamics. Notably, 50% or more of clinically relevant amino acid substitutions map onto the identified signal transduction pathways. This suggests that clinically important variation may affect, or be driven by, differences in allosteric behavior, providing a mechanism by which amino acid substitutions may affect the relationship between spectrum of activity, catalytic turnover, and potential allosteric behavior in this clinically important enzyme family. Simulations of the type presented here will help in identifying and analyzing such differences.

Antibiotics are crucial drugs for treating and preventing bacterial infections, but some bacteria are evolving ways to resist their effects. This 'antibiotic resistance' threatens lives and livelihoods worldwide. ß-lactam antibiotics, like penicillin, are some of the most commonly used, but some bacteria can now make enzymes called ß-lactamases, which destroy these antibiotics. Dozens of different types of ß-lactamases now exist, each with different properties. Two of the most medically important are TEM-1 and KPC-2. One way to counteract ß-lactamases is with drugs called inhibitors that stop the activity of these enzymes. The approved ß-lactamase inhibitors work by blocking the part of the enzyme that binds and destroys antibiotics, known as the 'active site'. The ß-lactamases have evolved, some of which have the ability to resist the effects of known inhibitors. It is possible that targeting parts of ß-lactamases far from the active site, known as 'allosteric sites', might get around these new bacterial defences. A molecule that binds to an allosteric site might alter the enzyme's shape, or restrict its movement, making it unable to do its job. Galdadas, Qu et al. used simulations to understand how molecules binding at allosteric sites affect enzyme movement. The experiments examined the structures of both TEM-1 and KPC-2, looking at how their shapes changed as molecules were removed from the allosteric site. This revealed how the allosteric sites and the active site are linked together. When molecules were taken out of the allosteric sites, they triggered ripples of shape change that travelled via loop-like structures across the surface of the enzyme. These loops contain over half of the known differences between the different types of ß-lactamases, suggesting mutations here may be responsible for changing which antibiotics each enzyme can destroy. In other words, changes in the 'ripples' may be related to the ability of the enzymes to resist particular antibiotics. Understanding how changes in one part of a ß-lactamase enzyme reach the active site could help in the design of new inhibitors. It might also help to explain how ß-lactamases evolve new properties. Further work could show why different enzymes are more or less active against different antibiotics.

A General Mechanism for Signal Propagation in the Nicotinic Acetylcholine Receptor Family.

Nicotinic acetylcholine receptors (nAChRs) modulate synaptic activity in the central nervous system. The α7 subtype, in particular, has attracted considerable interest in drug discovery as a target for several conditions, including Alzheimer's disease and schizophrenia. Identifying agonist-induced structural changes underlying nAChR activation is fundamentally important for understanding biological function and rational drug design. Here, extensive equilibrium and nonequilibrium molecular dynamics simulations, enabled by cloud-based high-performance computing, reveal the molecular mechanism by which structural changes induced by agonist unbinding are transmitted within the human α7 nAChR. The simulations reveal the sequence of coupled structural changes involved in driving conformational change responsible for biological function. Comparison with simulations of the α4ß2 nAChR subtype identifies features of the dynamical architecture common to both receptors, suggesting a general structural mechanism for signal propagation in this important family of receptors.