Gadegaardlindgaard3755

In redox metalloenzymes, the process of electron transfer often involves the concerted movement of a proton. These processes are referred to as proton-coupled electron transfer, and they underpin a wide variety of biological processes, including respiration, energy conversion, photosynthesis, and metalloenzyme catalysis. The mechanisms of proton delivery are incompletely understood, in part due to an absence of information on exact proton locations and hydrogen bonding structures in a bona fide metalloenzyme proton pathway. Here, we present a 2.1-Å neutron crystal structure of the complex formed between a redox metalloenzyme (ascorbate peroxidase) and its reducing substrate (ascorbate). In the neutron structure of the complex, the protonation states of the electron/proton donor (ascorbate) and all of the residues involved in the electron/proton transfer pathway are directly observed. This information sheds light on possible proton movements during heme-catalyzed oxygen activation, as well as on ascorbate oxidation.Tol-Pal is a multiprotein system present in the envelope of Gram-negative bacteria. Inactivation of this widely conserved machinery compromises the outer membrane (OM) layer of these organisms, resulting in hypersensitivity to many antibiotics. Mutants in the tol-pal locus fail to complete division and form cell chains. This phenotype along with the localization of Tol-Pal components to the cytokinetic ring in Escherichia coli has led to the proposal that the primary function of the system is to promote OM constriction during division. Accordingly, a poorly constricted OM is believed to link the cell chains formed upon Tol-Pal inactivation. However, we show here that cell chains of E. coli tol-pal mutants are connected by an incompletely processed peptidoglycan (PG) layer. Genetic suppressors of this defect were isolated and found to overproduce OM lipoproteins capable of cleaving the glycan strands of PG. Among the factors promoting cell separation in mutant cells was a protein of previously unknown function (YddW), which we have identified as a divisome-localized glycosyl hydrolase that cleaves peptide-free PG glycans. Overall, our results indicate that the cell chaining defect of Tol-Pal mutants cannot simply be interpreted as a defect in OM constriction. Rather, the complex also appears to be required for the activity of several OM-localized enzymes with cell wall remodeling activity. Thus, the Tol-Pal system may play a more general role in coordinating OM invagination with PG remodeling at the division site than previously appreciated.Intestinal bile acids are known to modulate the germination and growth of Clostridioides difficile Here we describe a role for intestinal bile acids in directly binding and neutralizing TcdB toxin, the primary determinant of C. difficile disease. We show that individual primary and secondary bile acids reversibly bind and inhibit TcdB to varying degrees through a mechanism that requires the combined oligopeptide repeats region to which no function has previously been ascribed. We find that bile acids induce TcdB into a compact "balled up" conformation that is no longer able to bind cell surface receptors. Lastly, through a high-throughput screen designed to identify bile acid mimetics we uncovered nonsteroidal small molecule scaffolds that bind and inhibit TcdB through a bile acid-like mechanism. In addition to suggesting a role for bile acids in C. difficile pathogenesis, these findings provide a framework for development of a mechanistic class of C. difficile antitoxins.Conservation of our cultural heritage is fundamental for conveying to future generations our culture, traditions, and ways of thinking and behaving. Cleaning art, in particular modern/contemporary paintings, with traditional tools could be risky and impractical, particularly on large collections of important works to be transferred to future generations. We report on advanced cleaning systems, based on twin-chain polymer networks made of poly(vinyl alcohol) (PVA) chains, semiinterpenetrated (semi-IPN) with PVA of lower molecular weight (L-PVA). Interpenetrating L-PVA causes a change from gels with oriented channels to sponge-like semi-IPNs with disordered interconnected pores, conferring different gel (and solvent) dynamics. These features grant residue-free, time efficient cleaning capacity and effective dirt capture, defeating risks for the artifact, making possible a safer treatment of important collections, unconceivable with conventional methods. We report as an example the conservation of Jackson Pollock's masterpieces, cleaned in a controlled way, safety and selectivity with unprecedented performance. Copyright © 2020 the Author(s). Published by PNAS.The mitochondria of various tissues from mice, naked mole rats (NMRs), and bats possess two mechanistically similar systems to prevent the generation of mitochondrial reactive oxygen species (mROS) hexokinases I and II and creatine kinase bound to mitochondrial membranes. Both systems operate in a manner such that one of the kinase substrates (mitochondrial ATP) is electrophoretically transported by the ATP/ADP antiporter to the catalytic site of bound hexokinase or bound creatine kinase without ATP dilution in the cytosol. One of the kinase reaction products, ADP, is transported back to the mitochondrial matrix via the antiporter, again through an electrophoretic process without cytosol dilution. https://www.selleckchem.com/products/actinomycin-d.html The system in question continuously supports H+-ATP synthase with ADP until glucose or creatine is available. Under these conditions, the membrane potential, ∆ψ, is maintained at a lower than maximal level (i.e., mild depolarization of mitochondria). This ∆ψ decrease is sufficient to completely inhibit mROS generation. In 2.5-y-old mice, mild depolarization disappears in the skeletal muscles, diaphragm, heart, spleen, and brain and partially in the lung and kidney. This age-dependent decrease in the levels of bound kinases is not observed in NMRs and bats for many years. As a result, ROS-mediated protein damage, which is substantial during the aging of short-lived mice, is stabilized at low levels during the aging of long-lived NMRs and bats. It is suggested that this mitochondrial mild depolarization is a crucial component of the mitochondrial anti-aging system. Copyright © 2020 the Author(s). Published by PNAS.