BackgroundOctopamine receptors (OARs) perform key functions in the biological pathways of primarily invertebrates, making this class of G-protein coupled receptors (GPCRs) a potentially good target for insecticides. However, the lack of structural and experimental data for this insect-essential GPCR family has promoted the development of homology models that are good representations of their biological equivalents for in silico screening of small molecules.MethodsTwo Anopheles gambiae OARs were cloned, analysed and functionally characterized using a heterologous cell reporter system. Four antagonist- and four agonist-binding homology models were generated and virtually screened by docking against compounds obtained from the ZINC database. Resulting compounds from the virtual screen were tested experimentally using an in vitro reporter assay and in a mosquito larvicide bioassay.ResultsSix An. gambiae OAR/tyramine receptor genes were identified. Phylogenetic analysis revealed that the OAR (AGAP000045) that encodes two open reading frames is an α-adrenergic-like receptor. Both splice variants signal through cAMP and calcium. Mutagenesis analysis revealed that D100 in the TM3 region and S206 and S210 in the TM5 region are important to the activation of the GPCR. Some 2,150 compounds from the virtual screen were structurally analysed and 70 compounds were experimentally tested against AgOAR45B expressed in the GloResponse™CRE-luc2P HEK293 reporter cell line, revealing 21 antagonists, 17 weak antagonists, 2 agonists, and 5 weak agonists.ConclusionReported here is the functional characterization of two An. gambiae OARs and the discovery of new OAR agonists and antagonists based on virtual screening and molecular dynamics simulations. Four compounds were identified that had activity in a mosquito larva bioassay, three of which are imidazole derivatives. This combined computational and experimental approach is appropriate for the discovery of new and effective insecticides.Electronic supplementary materialThe online version of this article (doi:10.1186/1475-2875-13-434) contains supplementary material, which is available to authorized users.
Insulin plays a central role in the regulation of metabolism in humans. Mutations in the insulin gene can impair the folding of its precursor protein, proinsulin, and cause permanent neonatal-onset diabetes mellitus known as Mutant INS-gene induced Diabetes of Youth (MIDY) with insulin deficiency. To gain insights into the molecular basis of this diabetes-associated mutation, we perform molecular dynamics simulations in wild-type and mutant (CysA7 to Tyr or C(A7)Y) insulin A chain in aqueous solutions. The C(A7)Y mutation is one of the identified mutations that impairs the protein folding by substituting the cysteine residue which is required for the disulfide bond formation. A comparative analysis reveals structural differences between the wild-type and the mutant conformations. The analyzed mutant insulin A chain forms a metastable state with major effects on its N-terminal region. This suggests that MIDY mutant involves formation of a partially folded intermediate with conformational change in N-terminal region in A chain that generates flexible N-terminal domain. This may lead to the abnormal interactions with other proinsulins in the aggregation process.
Octopamine receptors (OARs) perform key biological functions in invertebrates, making this class of G-protein coupled receptors (GPCRs) worth considering for insecticide development. However, no crystal structures and very little research exists for OARs. Furthermore, GPCRs are large proteins, are suspended in a lipid bilayer, and are activated on the millisecond timescale, all of which make conventional molecular dynamics (MD) simulations infeasible, even if run on large supercomputers. However, accelerated Molecular Dynamics (aMD) simulations can reduce this timescale to even hundreds of nanoseconds, while running the simulations on graphics processing units (GPUs) would enable even small clusters of GPUs to have processing power equivalent to hundreds of CPUs. Our results show that aMD simulations run on GPUs can successfully obtain the active and inactive state conformations of a GPCR on this reduced timescale. Furthermore, we discovered a potential alternate active-state agonist-binding position in the octopamine receptor which has yet to be observed and may be a novel GPCR agonist-binding position. These results demonstrate that a complex biological system with an activation process on the millisecond timescale can be successfully simulated on the nanosecond timescale using a simple computing system consisting of a small number of GPUs. Proteins 2016; 84:1480-1489. © 2016 Wiley Periodicals, Inc.
Digital twins of product and production processes are being used more and more frequently. The reason for this are the benefits for all departments involved. Therefore, it is not only important to share an IT platform as a 3D collaboration environment with uniform data structures, so all employees can work with the same database. As well it is important to give all interdisciplinary departments the opportunity to work in a common simulation and interact with it as a digital twin of the product and production process. Each department must be enabled to add their information, to collaboratively design a working digital twin of the real product. In addition, it must be possible for several people to work together simultaneously in a shared virtual environment. Voice chat, video chat and graphic simulation features such as virtual and augmented reality must be integrated to enable collaboration and interaction with the digital twin. Another challenge is to allow multiple participants to take part in these virtual meetings simultaneously, even if they use different end devices. The paper presents a 3D collaboration environment developed in research projects based on the development environment for graphic simulations Unity3D, with the ability to include various user concepts and functions for the specialist departments and allows access for the employees as a multi-user application on different end devices.
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