Total In Vitro Biosynthesis of the Thioamitide Thioholgamide and Investigation of the Pathway

Thioholgamides are ribosomally synthesized and post-translationally modified peptides (RiPPs) with potent activity against cancerous cell lines and an unprecedented structure. Despite being one of the most structurally and chemically complex RiPPs, very few biosynthetic steps have been elucidated. Here, we report the complete in vitro reconstitution of the biosynthetic pathway. We demonstrate that thioamidation is the first step and acts as a gatekeeper for downstream processing. Thr dehydration follows thioamidation, and our studies reveal that both these modifications require the formation of protein complexes – ThoH/I and ThoC/D. Harnessing the power of AlphaFold we deduce that ThoD acts as a lyase and also propose putative catalytic residues. ThoF catalyzes the oxidative decarboxylation of the terminal Cys and the subsequent macrocyclization is facilitated by ThoE. This is followed by Ser dehydration, which is also carried out by ThoC/D. ThoG is responsible for histidine bis-N-methylation, which is a prerequisite for His β-hydroxylation – a modification carried out by ThoJ. The last step of the pathway is the removal of the leader peptide by ThoK to afford mature thioholgamide.

Explaining Valence Asymmetries in Value Learning: A Reinforcement Learning Account

To understand how acquired value impacts how we perceive and process stimuli, psychologists have developed the Value Learning Task (VLT; e.g., Raymond & O’Brien, 2009). The task consists of a series of trials in which participants attempt to maximize accumulated winnings as they make choices from a pair of presented images associated with probabilistic win, loss, or no-change outcomes. Despite the task having a symmetric outcome structure for win and loss pairs, people learn win associations better than loss associations (Lin, Cabrera-Haro, & Reuter-Lorenz, 2020). This asymmetry could lead to differences when the stimuli are probed in subsequent tasks, compromising inferences about how acquired value affects downstream processing. We investigate the nature of the asymmetry using a standard error-driven reinforcement learning model with a softmax choice rule. Despite having no special role for valence, the model yields the asymmetry observed in human behavior, whether the model parameters are set to maximize empirical fit, or task payoff. The asymmetry arises from an interaction between a neutral initial value estimate and a choice policy that exploits while exploring, leading to more poorly discriminated value estimates for loss stimuli. We also show how differences in estimated individual learning rates help to explain individual differences in the observed win-loss asymmetries, and how the final value estimates produced by the model provide a simple account of a post-learning explicit value categorization task.

Wood Ash Based Treatment of Anaerobic Digestate: State-of-the-Art and Possibilities

The problem of current agricultural practices is not limited to land management but also to the unsustainable consumption of essential nutrients for plants, such as phosphorus. This article focuses on the valorization of wood ash and anaerobic digestate for the preparation of a slow-release fertilizer. The underlying chemistry of the blend of these two materials is elucidated by analyzing the applications of the mixture. First, the feasibility of employing low doses (≤1 g total solids (TS) ash/g TS digestate) of wood ash is explained as a way to improve the composition of the feedstock of anaerobic digestion and enhance biogas production. Secondly, a detailed description concerning high doses of wood ash and their uses in the downstream processing of the anaerobic digestate to further enhance its stability is offered. Among all the physico-chemical phenomena involved, sorption processes are meticulously depicted, since they are responsible for nutrient recovery, dewatering, and self-hardening in preparing a granular fertilizer. Simple activation procedures (e.g., carbonization, carbonation, calcination, acidification, wash, milling, and sieving) are proposed to promote immobilization of the nutrients. Due to the limited information on the combined processing of wood ash and the anaerobic digestate, transformations of similar residues are additionally considered. Considering all the possible synergies in the anaerobic digestion and the downstream stages, a dose of ash of 5 g TS ash/g TS digestate is proposed for future experiments.

Enhancing Downstream Processing Of Algal-Based Biofuel Using Novel Fused Bacteria And Green Solvents

The present study investigated the utilization of algal biomass to produce bio-oil and acetone, butanol, and ethanol (ABE) products. Novel Clostridia fusants (C. beijernickii + C. thermocellum-CbCt and C. acetobutylicum + C. thermocellocum-CaCt) were developed using protoplast fusion technique and subsequently subjected to UV radiation for strain enhancement. Resultant mutated fusants showed improvement in thermal stability and higher resistance to biobutanol toxicity. Algal biomass was initially subjected to various hydrolysis treatments prior to fermentation. Combination treatment of thermal, chemical, and enzymatic resulted in maximum sugar release of 27.78 g/L. Maximum biobutanol concentration from fermentation using CbCt resulted in 7.98 g/L. Fermentation using CaCt produced a concentration of 7.39 g/L. Oil extraction from virgin algae investigated a green, bio-based approach using terpenes with ultrasonication and a modified, Bligh and Dyer method, separately. Combination method, ultrasonication followed by the modified Bligh and Dyer, resulted in oil yield of 46.27% (dlimonene) and 39.85% (p-cymene). Oil extraction was also produced from an algae sample following fermentation. Combined extraction method using fermentation sample resulted in oil yield of 65.04%.

Enhancing Downstream Processing Of Algal-Based Biofuel Using Novel Fused Bacteria And Green Solvents

The present study investigated the utilization of algal biomass to produce bio-oil and acetone, butanol, and ethanol (ABE) products. Novel Clostridia fusants (C. beijernickii + C. thermocellum-CbCt and C. acetobutylicum + C. thermocellocum-CaCt) were developed using protoplast fusion technique and subsequently subjected to UV radiation for strain enhancement. Resultant mutated fusants showed improvement in thermal stability and higher resistance to biobutanol toxicity. Algal biomass was initially subjected to various hydrolysis treatments prior to fermentation. Combination treatment of thermal, chemical, and enzymatic resulted in maximum sugar release of 27.78 g/L. Maximum biobutanol concentration from fermentation using CbCt resulted in 7.98 g/L. Fermentation using CaCt produced a concentration of 7.39 g/L. Oil extraction from virgin algae investigated a green, bio-based approach using terpenes with ultrasonication and a modified, Bligh and Dyer method, separately. Combination method, ultrasonication followed by the modified Bligh and Dyer, resulted in oil yield of 46.27% (dlimonene) and 39.85% (p-cymene). Oil extraction was also produced from an algae sample following fermentation. Combined extraction method using fermentation sample resulted in oil yield of 65.04%.

All-in-one Superparamagnetic and SERS-active Niosomes for Dual-targeted in Vitro Detection of Breast Cancer Cells

BackgroundIn vitro and in vivo biosensing through surface-enhanced Raman scattering often suffer from signal contamination diminishing both the limit of detection and quantification. However, overcoming the lack of specificity requires excessive nanoparticle concentrations, which may lead to adverse side effects if applied to patients. ResultsWe propose encapsulation of iron oxide (FexOy) and gold (Au) nanoparticles (NPs) into the bilayer structure of transferrin-modified niosomes. This approach enables achieving greatly enhanced and contamination-free SERS-signals in vitro as well as a dual-targeting functionality towards MCF-7 breast cancer cells. An in-depth characterization of FexOyNPs- and AuNPs-loaded niosomes (AuNPs/FexOyNPs/NIO) after magnetic downstream processing reveals defined hybrid niosome structures, which show a long-term SERS-signal stability in various media such as MCF-7 cell culture medium. In vitro 2D-SERS imaging unveil a successful incorporation of a non-toxic dose of hybrid NPs into MCF-7 cells, which leads to strong and almost contamination-free SERS-signals. The measured signal-to-noise ratio of the in vitro signal exceeds the values required by DIN 32645 for the successful validation of a detection method. ConclusionsThe hybrid niosomes can be considered a promising and efficient agent for the establishment and commercialization of a highly sensitive detection kit for monitoring cancerous tissue.

Nanovaccine Delivery Approaches and Advanced Delivery Systems for the Prevention of Viral Infections: From Development to Clinical Application

Viral infections causing pandemics and chronic diseases are the main culprits implicated in devastating global clinical and socioeconomic impacts, as clearly manifested during the current COVID-19 pandemic. Immunoprophylaxis via mass immunisation with vaccines has been shown to be an efficient strategy to control such viral infections, with the successful and recently accelerated development of different types of vaccines, thanks to the advanced biotechnological techniques involved in the upstream and downstream processing of these products. However, there is still much work to be done for the improvement of efficacy and safety when it comes to the choice of delivery systems, formulations, dosage form and route of administration, which are not only crucial for immunisation effectiveness, but also for vaccine stability, dose frequency, patient convenience and logistics for mass immunisation. In this review, we discuss the main vaccine delivery systems and associated challenges, as well as the recent success in developing nanomaterials-based and advanced delivery systems to tackle these challenges. Manufacturing and regulatory requirements for the development of these systems for successful clinical and marketing authorisation were also considered. Here, we comprehensively review nanovaccines from development to clinical application, which will be relevant to vaccine developers, regulators, and clinicians.