Engineering the Melanocortin-4 Receptor to Control Constitutive and Ligand-Mediated Gs Signaling In Vivo

The molecular and functional diversity of G protein–coupled receptors is essential to many physiological processes. However, this diversity presents a significant challenge to understanding the G protein–mediated signaling events that underlie a specific physiological response. To increase our understanding of these processes, we sought to gain control of the timing and specificity of Gs signaling in vivo. We used naturally occurring human mutations to develop two Gs-coupled engineered receptors that respond solely to a synthetic ligand (RASSLs). Our Gs-coupled RASSLs are based on the melanocortin-4 receptor, a centrally expressed receptor that plays an important role in the regulation of body weight. These RASSLs are not activated by the endogenous hormone α-melanocyte-stimulating hormone but respond potently to a selective synthetic ligand, tetrahydroisoquinoline. The RASSL variants reported here differ in their intrinsic basal activities, allowing the separation of the effects of basal signaling from ligand-mediated activation of the Gs pathway in vivo. These RASSLs can be used to activate Gs signaling in any tissue, but would be particularly useful for analyzing downstream events that mediate body weight regulation in mice. Our study also demonstrates the use of human genetic variation for protein engineering

Application of agent-based modelling to simulate ribosome translation

Translation is a key process in the cell that encompasses the formation of proteins. However, how the translation mechanisms are affected by physiological changes is yet to be determined. Saccharomyces cerevisiae is one of the most used microorganisms to express recombinant proteins, showing great industrial/commercial value. Modelling the translation process in this yeast can thus bring forward novel insights into its mechanisms and how they are affected by changes in the environment. The present work introduces an agent-based model describing the elongation step of the translation process in the yeast. The simulated and theoretical elongation times were almost identical, with a standard deviation of 0.0018\\%, demonstrating the usefulness of the model to simulate this type of scenarios. Results also show a negative correlation between tRNA levels and estimated decoding times of codons, in accordance with biological knowledge. The model holds considerable potential to help unveil new ways of manipulation and thus increase the production of economically relevant yeast-derived products, namely biopharmaceuticals. Further development will address more complex scenarios, such as ribosome queuing or all the phases in the translation process.This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit, BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte, and of the PhD Grant SFRH/BD/143491/2019. Additionally, it received funding through Base Funding - UIDB/00511/2020 of the Laboratory for Process Engineering, Environment, Biotechnology and Energy – LEPABE - funded by national funds through the FCT/MCTES (PIDDAC).info:eu-repo/semantics/publishedVersio