A versatile disulfide-driven recycling system for NADP+ with high cofactor turnover number

NADP+-dependent enzymes are important in many biocatalytic processes to generate high-value chemicals for the pharmaceutical and food industry; hence, a costeffective, efficient, and environmentally friendly recycling system for the relatively expensive and only marginally stable enzyme cofactor NADP+ offers significant benefits. NADP+ regeneration schemes have previously been described, but their application is severely limited by the low total turnover numbers (TTN) for the cofactor. Here, we report a glutathione-based recycling system that combines glutaredoxin from E. coli (EcGRX) and the glutathione reductase from S. cerevisiae (ScGR) for NADP+ regeneration. This system employs inexpensive latent organic disulfides such as oxidized cysteine or 2-hydroxyethyl disulfide (HED) as oxidizing agents and allows NADP+ recycling under both aerobic and anaerobic conditions with a TTN in excess of 5 × 105, indicating that each regeneration cycle is 99.9998% selective toward forming the cofactor. Accordingly, for each 1 mol of product generated, less than $0.05 of cofactor is needed. Finally, the EcGRX/ScGR pair is compatible with eight enzymes in the guanosine monophosphate (GMP) biosynthetic pathway, giving the corresponding isotopically labeled nucleotide in high yield. The glutathione-based NADP+ recycling system has potential for biocatalytic applications in academic and industrial settings

A scaffold replacement approach towards new sirtuin 2 inhibitors

Sirtuins (SIRT1-SIRT7) are an evolutionary conserved family of NAD(+)-dependent protein deacylases regulating the acylation state of epsilon-N-lysine residues of proteins thereby controlling key biological processes. Numerous studies have found association of the aberrant enzymatic activity of SIRTs with various diseases like diabetes, cancer and neurodegenerative disorders. Previously, we have shown that substituted 2-alkyl-chroman-4-one/chromone derivatives can serve as selective inhibitors of SIRT2 possessing an antiproliferative effect in two human cancer cell lines. In this study, we have explored the bioisosteric replacement of the chroman-4-one/chromone core structure with different less lipophilic bicyclic scaffolds to overcome problems associated to poor physiochemical properties due to a highly lipophilic substitution pattern required for achieve a good inhibitory effect. Various new derivatives based on the quinolin-4(1H)-one scaffold, bicyclic secondary sulfonamides or saccharins were synthesized and evaluated for their SIRT inhibitory effect. Among the evaluated scaffolds, the benzothiadiazine-1,1-dioxide-based compounds showed the highest SIRT2 inhibitory activity. Molecular modeling studies gave insight into the binding mode of the new scaffold-replacement analogues.Peer reviewe

A Neptune Orbiter Concept Using Drag Modulated Aerocaptue (DMA) and the Adaptable, Deployable Entry and Placement Technology (ADEPT)

Conceptual Neptune orbiter was designed for the purpose of assessing mission feasibilityBuilt off of the 2017 Pre-Decadal Study, but adapted for drag modulation aerocapture.Science payload includes: Narrow Angle camera, Doppler Imager, Magnetometer, Atmospheric Probe (w/ ASI, Nephelometer, Mass Spectrometer). Baseline concept of operations releases probe prior to orbit insertion, but investigations are ongoing to assess the feasibility of bringing the probe to orbit before release

Epigenetic regulation of PPARGC1A in human type 2 diabetic islets and effect on insulin secretion

Aims/hypothesis Insulin secretion in pancreatic islets is dependent upon mitochondrial function and production of ATP. The transcriptional coactivator peroxisome proliferator activated receptor gamma coactivator-1 alpha (protein PGC-1 alpha; gene PPARGC1A) is a master regulator of mitochondrial genes and its expression is decreased and related to impaired oxidative phosphorylation in muscle from patients with type 2 diabetes. Whether it plays a similar role in human pancreatic islets is not known. We therefore investigated if PPARGC1A expression is altered in islets from patients with type 2 diabetes and whether this expression is influenced by genetic (PPARGC1A Gly482Ser polymorphism) and epigenetic (DNA methylation) factors. We also tested if experimental downregulation of PPARGC1A expression in human islets influenced insulin secretion. Methods The PPARGC1A Gly482Ser polymorphism was genotyped in human pancreatic islets from 48 non-diabetic and 12 type 2 diabetic multi-organ donors and related to PPARGC1A mRNA expression. DNA methylation of the PPARGC1A promoter was analysed in pancreatic islets from ten type 2 diabetic and nine control donors. Isolated human islets were transfected with PPARGC1A silencing RNA (siRNA). Results PPARGC1A mRNA expression was reduced by 90% (p < 0.005) and correlated with the reduction in insulin secretion in islets from patients with type 2 diabetes. After downregulation of PPARGC1A expression in human islets by siRNA, insulin secretion was reduced by 41% (p <= 0. 01). We were able to ascribe reduced PPARGC1A expression in islets to both genetic and epigenetic factors, i.e. a common PPARGC1A Gly482Ser polymorphism was associated with reduced PPARGC1A mRNA expression (p < 0.00005) and reduced insulin secretion (p < 0.05). In support of an epigenetic influence, the PPARGC1A gene promoter showed a twofold increase in DNA methylation in diabetic islets compared with non-diabetic islets (p < 0.04). Conclusions/Interpretation We have shown for the first time that PPARGC1A might be important in human islet insulin secretion and that expression of PPARGC1A in human islets can be regulated by both genetic and epigenetic factors

Genomic organisation and alternative splicing of mouse and human thioredoxin reductase 1 genes

BACKGROUND: Thioredoxin reductase (TR) is a redox active protein involved in many cellular processes as part of the thioredoxin system. Presently there are three recognised forms of mammalian thioredoxin reductase designated as TR1, TR3 and TGR, that represent the cytosolic, mitochondrial and novel forms respectively. In this study we elucidated the genomic organisation of the mouse (Txnrd1) and human thioredoxin reductase 1 genes (TXNRD1) through library screening, restriction mapping and database mining. RESULTS: The human TXNRD1 gene spans 100 kb of genomic DNA organised into 16 exons and the mouse Txnrd1 gene has a similar exon/intron arrangement. We also analysed the alternative splicing patterns displayed by the mouse and human thioredoxin reductase 1 genes and mapped the different mRNA isoforms with respect to genomic organisation. These isoforms differ at the 5' end and encode putative proteins of different molecular mass. Genomic DNA sequences upstream of mouse exon 1 were compared to the human promoter to identify conserved elements. CONCLUSIONS: The human and mouse thioredoxin reductase 1 gene organisation is highly conserved and both genes exhibit alternative splicing at the 5' end. The mouse and human promoters share some conserved sequences

Knockdown of Cytosolic Glutaredoxin 1 Leads to Loss of Mitochondrial Membrane Potential: Implication in Neurodegenerative Diseases

Mitochondrial dysfunction including that caused by oxidative stress has been implicated in the pathogenesis of neurodegenerative diseases. Glutaredoxin 1 (Grx1), a cytosolic thiol disulfide oxido-reductase, reduces glutathionylated proteins to protein thiols and helps maintain redox status of proteins during oxidative stress. Grx1 downregulation aggravates mitochondrial dysfunction in animal models of neurodegenerative diseases, such as Parkinson's and motor neuron disease. We examined the mechanism underlying the regulation of mitochondrial function by Grx1. Downregulation of Grx1 by shRNA results in loss of mitochondrial membrane potential (MMP), which is prevented by the thiol antioxidant, α-lipoic acid, or by cyclosporine A, an inhibitor of mitochondrial permeability transition. The thiol groups of voltage dependent anion channel (VDAC), an outer membrane protein in mitochondria but not adenosine nucleotide translocase (ANT), an inner membrane protein, are oxidized when Grx1 is downregulated. We then examined the effect of β-N-oxalyl amino-L-alanine (L-BOAA), an excitatory amino acid implicated in neurolathyrism (a type of motor neuron disease), that causes mitochondrial dysfunction. Exposure of cells to L-BOAA resulted in loss of MMP, which was prevented by overexpression of Grx1. Grx1 expression is regulated by estrogen in the CNS and treatment of SH-SY5Y cells with estrogen upregulated Grx1 and protected from L-BOAA mediated MMP loss. Our studies demonstrate that Grx1, a cytosolic oxido-reductase, helps maintain mitochondrial integrity and prevents MMP loss caused by oxidative insult. Further, downregulation of Grx1 leads to mitochondrial dysfunction through oxidative modification of the outer membrane protein, VDAC, providing support for the critical role of Grx1 in maintenance of MMP

The aquaculture supply chain in the time of covid-19 pandemic: vulnerability, resilience, solutions and priorities at the global scale

The COVID-19 global pandemic has had severe, unpredictable and synchronous impacts on all levels of perishable food supply chains (PFSC), across multiple sectors and spatial scales. Aquaculture plays a vital and rapidly expanding role in food security, in some cases overtaking wild caught fisheries in the production of high-quality animal protein in this PFSC. We performed a rapid global assessment to evaluate the effects of the COVID-19 pandemic and related emerging control measures on the aquaculture supply chain. Socio-economic effects of the pandemic were analysed by surveying the perceptions of stakeholders, who were asked to describe potential supply-side disruption, vulnerabilities and resilience patterns along the production pipeline with four main supply chain components: a) hatchery, b) production/processing, c) distribution/logistics and d) market. We also assessed different farming strategies, comparing land- vs. sea-based systems; extensive vs. intensive methods; and with and without integrated multi-trophic aquaculture, IMTA. In addition to evaluating levels and sources of economic distress, interviewees were asked to identify mitigation solutions adopted at local / internal (i.e., farm-site) scales, and to express their preference on national / external scale mitigation measures among a set of a priori options. Survey responses identified the potential causes of disruption, ripple effects, sources of food insecurity, and socio-economic conflicts. They also pointed to various levels of mitigation strategies. The collated evidence represents a first baseline useful to address future disaster-driven responses, to reinforce the resilience of the sector and to facilitate the design reconstruction plans and mitigation measures, such as financial aid strategies.publishe

The synergistic impacts of anthropogenic stressors and COVID-19 on aquaculture: a current global perspective

The rapid, global spread of COVID-19, and the measures intended to limit or slow its propagation, are having major impacts on diverse sectors of society. Notably, these impacts are occurring in the context of other anthropogenic-driven threats including global climate change. Both anthropogenic stressors and the COVID-19 pandemic represent significant economic challenges to aquaculture systems across the globe, threatening the supply chain of one of the most important sources of animal protein, with potential disproportionate impacts on vulnerable communities. A web survey was conducted in 47 countries in the midst of the COVID-19 pandemic to assess how aquaculture activities have been affected by the pandemic, and to explore how these impacts compare to those from climate change. A positive correlation between the effects of the two categories of drivers was detected, but analysis suggests that the pandemic and the anthropogenic stressors affect different parts of the supply chain. The immediate measurable reported losses varied with aquaculture typology (land vs. marine, and intensive vs. extensive). A comparably lower impact on farmers reporting the use of integrated multitrophic aquaculture (IMTA) methods suggests that IMTA might enhance resilience to multiple stressors by providing different market options under the COVID-19 pandemic. Results emphasize the importance of assessing detrimental effects of COVID-19 under a multiple stressor lens, focusing on areas that have already locally experienced economic loss due to anthropogenic stressors in the last decade. Holistic policies that simultaneously address other ongoing anthropogenic stressors, rather than focusing solely on the acute impacts of COVID-19, are needed to maximize the long-term resilience of the aquaculture sector.publishe

Dopaminergic D1 receptor signalling is necessary, but not sufficient for cued fear memory destabilisation

Rationale. Pharmacological targeting of memory reconsolidation is a promising therapeutic strategy for the treatment of fear memory-related disorders. However, the success of reconsolidation-based approaches depends upon the effective destabilisation of the fear memory by memory reactivation. Objectives. Here, we aimed to determine the functional involvement of dopamine D1 receptors in cued fear memory destabilisation, using systemic drug administration. Results. We observed that direct D1 receptor agonism was not sufficient to stimulate tone fear memory destabilisation to facilitate reconsolidation disruption by the glucocorticoid receptor antagonist mifepristone. Instead, administration of the nootropic nefiracetam did facilitate mifepristone-induced amnesia, in a manner that was dependent upon dopamine D1 receptor activation, although. Finally, while the combined treatment with nefiracetam and mifepristone did not confer fear-reducing effects under conditions of extinction learning, there was some evidence that mifepristone reduces fear expression irrespective of memory reactivation parameters. Conclusions. The use of combination pharmacological treatment to stimulate memory destabilisation and impair reconsolidation has potential therapeutic benefits, without risking a maladaptive increase of fear