Exploring the enzymatic repertoires of Bacteria and Archaea and their associations with metabolic maps.

The evolution, survival, and adaptation of microbes are consequences of gene duplication, acquisition, and divergence in response to environmental challenges. In this context, enzymes play a central role in the evolution of organisms, because they are fundamental in cell metabolism. Here, we analyzed the enzymatic repertoire in 6,467 microbial genomes, including their abundances, and their associations with metabolic maps. We found that the enzymes follow a power-law distribution, in relation to the genome sizes. Therefore, we evaluated the total proportion enzymatic classes in relation to the genomes, identifying a descending-order proportion: transferases (EC:2.-), hydrolases (EC:3.-), oxidoreductases (EC:1.-), ligases (EC:6.-), lyases (EC:4.-), isomerases (EC:5.-), and translocases (EC:7-.). In addition, we identified a preferential use of enzymatic classes in metabolism pathways for xenobiotics, cofactors and vitamins, carbohydrates, amino acids, glycans, and energy. Therefore, this analysis provides clues about the functional constraints associated with the enzymatic repertoire of functions in Bacteria and Archaea.

Mechanism of voltage sensing in Ca2+- and voltage-activated K+ (BK) channels.

In neurosecretion, allosteric communication between voltage sensors and Ca2+ binding in BK channels is crucially involved in damping excitatory stimuli. Nevertheless, the voltage-sensing mechanism of BK channels is still under debate. Here, based on gating current measurements, we demonstrate that two arginines in the transmembrane segment S4 (R210 and R213) function as the BK gating charges. Significantly, the energy landscape of the gating particles is electrostatically tuned by a network of salt bridges contained in the voltage sensor domain (VSD). Molecular dynamics simulations and proton transport experiments in the hyperpolarization-activated R210H mutant suggest that the electric field drops off within a narrow septum whose boundaries are defined by the gating charges. Unlike Kv channels, the charge movement in BK appears to be limited to a small displacement of the guanidinium moieties of R210 and R213, without significant movement of the S4.

Sequence and structural conservation reveal fingerprint residues in TRP channels.

Transient receptor potential (TRP) proteins are a large family of cation-selective channels, surpassed in variety only by voltage-gated potassium channels. Detailed molecular mechanisms governing how membrane voltage, ligand binding, or temperature can induce conformational changes promoting the open state in TRP channels are still a matter of debate. Aiming to unveil distinctive structural features common to the transmembrane domains within the TRP family, we performed phylogenetic reconstruction, sequence statistics, and structural analysis over a large set of TRP channel genes. Here, we report an exceptionally conserved set of residues. This fingerprint is composed of twelve residues localized at equivalent three-dimensional positions in TRP channels from the different subtypes. Moreover, these amino acids are arranged in three groups, connected by a set of aromatics located at the core of the transmembrane structure. We hypothesize that differences in the connectivity between these different groups of residues harbor the apparent differences in coupling strategies used by TRP subgroups.

Glucosamine-P and rhodococcal ADP-glucose pyrophosphorylases: A hint to (re)discover (actino)bacterial amino sugar metabolism.

Glycogen was described as a temporal storage molecule in rhodococci, interconnecting lipids and carbon availability. The Rhodococcus jostii ADP-glucose pyrophosphorylase (ADP-GlcPPase) kinetic and regulatory properties support this role. Curiously, the enzyme uses glucosamine-1P as alternative substrate. Herein, we report the in-depth study of glucosamine-1P activity and its regulation in two rhodocoocal ADP-GlcPPases, finding that glucosamine-6P (representing a metabolic carbon/nitrogen node) is a critical activator, then reinforcing the role of glycogen as an "intermediary metabolite" in rhodococci. Glucosamine-1P activity in rhodococcal ADP-GlcPPases responds to activation by metabolites improving their catalytic performance, strongly suggesting its metabolic feasibility. This work supports a scenario for new molecules/metabolites discovery and hypothesizes on evolutionary mechanisms underlying enzyme promiscuity opening novel metabolic features in (actino)bacteria.

Allosteric Modulators of Potential Targets Related to Alzheimer's Disease: a Review.

Among neurodegenerative disorders, Alzheimer's disease (AD) is the most common type of dementia, and there is an urgent need to discover new and efficacious forms of treatment for it. Pathological patterns of AD include cholinergic dysfunction, increased ß-amyloid (Aß) peptide concentration, the appearance of neurofibrillary tangles, among others, all of which are strongly associated with specific biological targets. Interactions observed between these targets and potential drug candidates in AD most often occur by competitive mechanisms driven by orthosteric ligands that sometimes result in the production of side effects. In this context, the allosteric mechanism represents a key strategy; this can be regarded as the selective modulation of such targets by allosteric modulators in an advantageous manner, as this may decrease the likelihood of side effects. The purpose of this review is to present an overview of compounds that act as allosteric modulators of the main biological targets related to AD.

A Model of Glucocorticoid Receptor Interaction With Coregulators Predicts Transcriptional Regulation of Target Genes.

Regulatory factors that control gene transcription in multicellular organisms are assembled in multicomponent complexes by combinatorial interactions. In this context, nuclear receptors provide well-characterized and physiologically relevant systems to study ligand-induced transcription resulting from the integration of cellular and genomic information in a cell- and gene-specific manner. Here, we developed a mathematical model describing the interactions between the glucocorticoid receptor (GR) and other components of a multifactorial regulatory complex controlling the transcription of GR-target genes, such as coregulator peptides. We support the validity of the model in relation to gene-specific GR transactivation with gene transcription data from A549 cells and in vitro real time quantification of coregulator-GR interactions. The model accurately describes and helps to interpret ligand-specific and gene-specific transcriptional regulation by the GR. The comprehensive character of the model allows future insight into the function and relative contribution of the molecular species proposed in ligand- and gene-specific transcriptional regulation.

Regulatory Properties of the ADP-Glucose Pyrophosphorylase from the Clostridial Firmicutes Member Ruminococcus albus.

ADP-glucose pyrophosphorylase from Firmicutes is encoded by two genes (glgC and glgD) leading to a heterotetrameric protein structure, unlike those in other bacterial phyla. The enzymes from two groups of Firmicutes, Bacillales and Lactobacillales, present dissimilar kinetic and regulatory properties. Nevertheless, no ADP-glucose pyrophosphorylase from Clostridiales, the third group in Firmicutes, has been characterized. For this reason, we cloned the glgC and glgD genes from Ruminococcus albus Different quaternary forms of the enzyme (GlgC, GlgD, and GlgC/GlgD) were purified to homogeneity and their kinetic parameters were analyzed. We observed that GlgD is an inactive monomer when expressed alone but increased the catalytic efficiency of the heterotetramer (GlgC/GlgD) compared to the homotetramer (GlgC). The heterotetramer is regulated by fructose-1,6-bisphosphate, phosphoenolpyruvate, and NAD(P)H. The first characterization of the Bacillales enzyme suggested that heterotetrameric ADP-glucose pyrophosphorylases from Firmicutes were unregulated. Our results, together with data from Lactobacillales, indicate that heterotetrameric Firmicutes enzymes are mostly regulated. Thus, the ADP-glucose pyrophosphorylase from Bacillales seems to have distinctive insensitivity to regulation.IMPORTANCE The enzymes involved in glycogen synthesis from Firmicutes have been less characterized in comparison with other bacterial groups. We performed kinetic and regulatory characterization of the ADP-glucose pyrophosphorylase from Ruminococcus albus Our results showed that this protein that belongs to different groups from Firmicutes (Bacillales, Lactobacillales, and Clostridiales) presents dissimilar features. This study contributes to the understanding of how this critical enzyme for glycogen biosynthesis is regulated in the Firmicutes group, whereby we propose that these heterotetrameric enzymes, with the exception of Bacillales, are allosterically regulated. Our results provide a better understanding of the evolutionary relationship of this enzyme family in Firmicutes.

Information flow and protein dynamics: the interplay between nuclear magnetic resonance spectroscopy and molecular dynamics simulations.

Proteins participate in information pathways in cells, both as links in the chain of signals, and as the ultimate effectors. Upon ligand binding, proteins undergo conformation and motion changes, which can be sensed by the following link in the chain of information. Nuclear magnetic resonance (NMR) spectroscopy and molecular dynamics (MD) simulations represent powerful tools for examining the time-dependent function of biological molecules. The recent advances in NMR and the availability of faster computers have opened the door to more detailed analyses of structure, dynamics, and interactions. Here we briefly describe the recent applications that allow NMR spectroscopy and MD simulations to offer unique insight into the basic motions that underlie information transfer within and between cells.